Difference between revisions of "Software2"

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This subset of programs was primarily written by Dr. Lifshitz.  It is really better to examine the full list of BIG software [[Software | here]].
 +
 
== Visualization ==
 
== Visualization ==
  
Line 8: Line 10:
 
== Simulation ==
 
== Simulation ==
 
===  analyze_simulation.pl  ===
 
===  analyze_simulation.pl  ===
 +
<nowiki>
  
 
This program takes one or more outputs of a simulation (e.g. output of run1 or run2)
 
This program takes one or more outputs of a simulation (e.g. output of run1 or run2)
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analyze_simulation.pl -verbose run1_cafl.i2i run1_ca.i2i > & sim.out
 
analyze_simulation.pl -verbose run1_cafl.i2i run1_ca.i2i > & sim.out
 
   
 
   
 +
</nowiki>
 
===  calc_diffusion3.pl  ===
 
===  calc_diffusion3.pl  ===
 +
<nowiki>
 
     This program reads in a 2D diffusion tracks and calculates a D for each file (track_\d+.out)
 
     This program reads in a 2D diffusion tracks and calculates a D for each file (track_\d+.out)
 
     by fitting a line to the MeanSquaredDisplacement vs time:
 
     by fitting a line to the MeanSquaredDisplacement vs time:
Line 75: Line 80:
  
 
   
 
   
 +
</nowiki>
 
===  compare_volume_measurements.pl  ===
 
===  compare_volume_measurements.pl  ===
 +
<nowiki>
  
 
This program combines data produced by cross_sections_of_voronoi (-vfile) with
 
This program combines data produced by cross_sections_of_voronoi (-vfile) with
Line 126: Line 133:
  
 
   
 
   
 +
</nowiki>
 
===  filter_beads.pl  ===
 
===  filter_beads.pl  ===
 +
<nowiki>
  
 
Reads in a file produced by makebeads2 (bead.info) and one produced by print_pixel_coords (mask.info).
 
Reads in a file produced by makebeads2 (bead.info) and one produced by print_pixel_coords (mask.info).
Line 172: Line 181:
  
 
   
 
   
 +
</nowiki>
 
===  graph_xpp2.pl  ===
 
===  graph_xpp2.pl  ===
 +
<nowiki>
  
 
Reads specified columns of data from an xpp output file.
 
Reads specified columns of data from an xpp output file.
Line 200: Line 211:
  
 
   
 
   
 +
</nowiki>
 
===  oxygen_krogh.pl  ===
 
===  oxygen_krogh.pl  ===
 +
<nowiki>
 
     NOT DONE YET.
 
     NOT DONE YET.
  
Line 215: Line 228:
  
 
   
 
   
 +
</nowiki>
 
===  random_path.pl  ===
 
===  random_path.pl  ===
 +
<nowiki>
  
 
Generate a random path (ie, diffusion).
 
Generate a random path (ie, diffusion).
Line 243: Line 258:
  
 
   
 
   
 +
</nowiki>
 
===  readbeads.pl  ===
 
===  readbeads.pl  ===
 +
<nowiki>
  
 
Reads in file produced by makebeads to check to see if sizes and amplitudes
 
Reads in file produced by makebeads to check to see if sizes and amplitudes
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 +
</nowiki>
 
===  run_random_path.pl  ===
 
===  run_random_path.pl  ===
 +
<nowiki>
  
 
Generate lots of random paths files (ie, diffusion) by calling random_path.pl multiple times..
 
Generate lots of random paths files (ie, diffusion) by calling random_path.pl multiple times..
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 +
</nowiki>
 
===  run_voronoi2.pl  ===
 
===  run_voronoi2.pl  ===
 +
<nowiki>
  
 
  This script calls cross_sections_of_voronoi many times, and produces an image and output text file from each run.
 
  This script calls cross_sections_of_voronoi many times, and produces an image and output text file from each run.
Line 312: Line 333:
  
 
   
 
   
 +
</nowiki>
 
===  voronoi_vs_fiji.pl  ===
 
===  voronoi_vs_fiji.pl  ===
 +
<nowiki>
 
compare the true volumes of voronoi regions (as reported by cross_sections_of_voronoi, e.g.  
 
compare the true volumes of voronoi regions (as reported by cross_sections_of_voronoi, e.g.  
 
voronoi_D170_0001_g0_G250_volume.txt) to those calculated
 
voronoi_D170_0001_g0_G250_volume.txt) to those calculated
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 +
</nowiki>
 
===  ball  ===
 
===  ball  ===
 +
<nowiki>
  
 
This program creates an elliptical shell of brightness the specified brightness (-i option)
 
This program creates an elliptical shell of brightness the specified brightness (-i option)
Line 382: Line 407:
 
         code in /home/lml/krypton/facil/ball.c  
 
         code in /home/lml/krypton/facil/ball.c  
 
   
 
   
 +
</nowiki>
 
===  calc_ca3  ===
 
===  calc_ca3  ===
 +
<nowiki>
 
#/home/lml/krypton/bin/calc_ca3  
 
#/home/lml/krypton/bin/calc_ca3  
  
Line 424: Line 451:
 
source code in ~lml/krypton/Walsh directory.
 
source code in ~lml/krypton/Walsh directory.
 
   
 
   
 +
</nowiki>
 
===  cellsim  ===
 
===  cellsim  ===
 +
<nowiki>
 
  [0]: in main, before MPI_Init()
 
  [0]: in main, before MPI_Init()
 
[0]: in main, after MPI_Init()
 
[0]: in main, after MPI_Init()
Line 625: Line 654:
 
       ~lml/vision/JSinger/src/channel              (steady state based upon Neher paper)
 
       ~lml/vision/JSinger/src/channel              (steady state based upon Neher paper)
  
−
===  cellsimL ===
+
</nowiki>
−
  [0]: in main, before MPI_Init()
+
===  channel_current2 ===
−
[0]: in main, after MPI_Init()
+
  <nowiki>
−
[0]: /home/lml/krypton/bin/cellsimL -h
+
channel_current2: like channel_current, but looks at [ca] over time at each BK (not just max [ca]).
−
help:
+
From that, it calculates the max BK current. Channel_current adds the max [ca] that each BK voxel sees
−
unable to parse command: -h
+
from each voxel in a Ryr cluster, to get the max [ca] that BK pixel sees.  But the max [ca] could occur
−
    This program simulates Ca and Na in a cylindrical cell with
+
at different times since the Ryr cluster has a spatial extent.  Experimental.
−
    cylindrical symmetry.  
+
 
−
    Note: It is recommended that the stderr output be examined for
+
Calculates the current which would be produced when each object in Ryr_image.i2i releases calcium which
−
          lines with the words: warning, Warning, error, or Error on them.
+
activates nearby BK channels in BK_image.i2i. Produces a histogram of these output currents.
−
Usage
+
The input images should have each pixel in an object having the object id as its value. These images can be
−
    cellsim [options] >& output_datafile
+
produced with "countobjs -Q" or "objs_via_ascent" 
 +
note:  a one pixel border around the image is ignored (added 12/8/09).
 +
This may take about one-half hour to run.
  
−
Options controlling the duration of the simulation:
+
Usage:
−
    -time #:       total time to simulate (in sec), default = 0.000000
+
channel_current [options] -f filename Ryr_image.i2i BK_image.i2i > im1_im2.hist
−
    -time_rate #: slow down time, ie take smaller dt steps during simulation.
+
Options:
−
                    this may be necessary if you are creating a large ca gradient,
+
-f filename resting: file containing [ca], in nM, as a function of distance and time. Right now this is
−
                    for example, with -ca_ring or -ca_src options (.5 means 1/2 the dt).
+
assumed to have 20 nm spacing and entries every msec. See
−
                    The number of iterations needed for convergence is
+
/home/lml/krypton/Zhuge/STIC/simulations/run32.ca as an example
−
                    roughly: I= (T*diff)/(dr*dr), where T is time, dr is
+
"resting" is the resting [ca], in nM, for the cell. It will we subtracted from the other
−
                    annulus thickness and diff is diffusion constant.
+
[ca] values to produce a "delta" [ca] which gets scaled linearly by Ryr intensity.
−
                    This ignores ion influx,etc. If you set diffusion rates very
+
The spark current used to produce this (see -C) should be roughly in the range expected
−
                    small, the iter # calculated will be too small if other stuff
+
for Ryr_image.i2i, since all delta [ca] is linearly extrapolated.
−
                    is happening, use -time_rate to compensate.
+
All distances > largest distance in filename will have their [ca] set to the [ca]
−
+
at the largest distance in filename.
−
Options controlling the geometry of the model:
+
note: Only the [ca] values at y == 1 are used. Is this what should be used?
−
    -radius #:    radius of the cell in microns, default = 6.000000
+
-u #: voltage (mV). BK conductance is a function of voltage. default = -80.000000
−
    -rings #:      radius of the cell in rings, default = 60
+
-U low delta hi: voltages from low to hi by delta. Max is 20 voltages
−
                    Ring thickness = cell radius/ # of rings.
+
-r #:         resolution (binsize) for output histogram in femtoAmps, default = 1000.000000
−
    -length #:    length of the cell in disks, default = 300
+
-R #: reveral potential of BK channel in mV, default = -80.000000
−
                    Each disk will be the same thickness as a ring.
 
−
    -bc Rmax Lmin Lmax:    boundary conditions at maximum radius and minimum and
 
−
                    maximum length coordinate (i.e., at the cell ends). These must
 
−
                    be the letters I (for infinite) or S (for solid, i.e. no ions go
 
−
                    through the membrane). The I option is not always accurate. default: S
 
−
    -mask image.i2i: only voxels in image.i2i > 0 are part of the cell.  Use this
 
−
                    to specify an arbitrary shape for the cell.
 
−
                    Not extensively tested.
 
−
    -spacing file: file specifies the spacing along each axis (i.e, size of each voxel).
 
−
                    spacing is in nm, one entry per voxel, comment lines ignored (#).
 
−
                    The cell direction (X,Y, or Z) must be the first letter on the line.
 
−
                    Other characters on the line will be ignored.
 
−
                    The number of entries must be < 2000 each direction.
 
−
                    Do not specify -rings,-radius,and -length if you use this option.
 
−
                    NOTE: the simulation time step goes as (length)^2, where length is
 
−
                          the length of the shortest side of all the voxels specified.
 
−
                    Any number of entries per line. First R, then D (length). Example:
 
−
                    # R direction (10 pixels in this direction, high res at low R)
 
−
                    X
 
−
                    25 50 75 100 200 300 400 500 600 1000
 
−
                    Y (lengthwise) direction (9 pixels in D, very high res in the middle)
 
−
                    100 200 300 400 500 540 550 560 1000
 
−
                    Use cyl2cart with -spacing option to convert output images to
 
−
                    a uniform spacing. (I haven't added that option yet)
 
−
                    Not extensively tested.
 
  
−
Options specifying substances (ions, buffers, compounds) present in the model (10 max):
+
  distances:
−
    -ion name free_concentration D valence:  
+
-w # # #: x,y, zweights for pixel distances (ie, pixel size), default = 1 for each.
−
                    concentration in mMolar, D in cm^2/sec, valence in absolute value.
+
-p #: pixel size in nm (i.e., what 1 means in -w option), default = 80.000000
−
                    valence is only used if -ion_src is also specified.
+
-m # :         maxdistance (in units used for -w, i.e., pixels) to look for the closest pixel, default = 10.000000
−
                    concentration is the initial free concentration.
+
 
−
    -buffer name total_concentration D:  
+
  which pixels or clusters to include:
−
                    concentration in mMolar, D in cm^2/sec.
+
-t # #: image1 and image2 thresholds, pixels > # are above threshold, default = 0.
−
                    concentration is the initial total concentration (sum of its free and all bound forms).
+
-e lowx lowy lowz hix hiy hiz:  only look at voxels within this extent (subvolume), 1-indexed, inclusive.
−
    -kin bufname ionname boundname onrate offrate D:  
+
-s minsize maxsize:     only look at objects in Ryr_image.i2i within the size range (inclusive, in voxels)
−
                    kinetics for interaction between bufname and ionname. Boundname is the
+
max size allowed = 100000
−
                    name to give the bound pair.  bufname and ionname must have been
+
-S minsize maxsize:     only look at objects in BK_image.i2i within the size range (inclusive, in voxels)
−
                    specified (with -ion -buffer or -kin options) prior to this option.
+
-i miniod maxiod:     only look at objects in Ryr_image.i2i within the iod range (inclusive. use -d also.)
−
                    onrate:in 1/(mMolar*seconds), offrate: in 1/seconds, D: in cm^2/sec 
+
-I miniod maxiod:     only look at objects in BK_image.i2i within the iod range (inclusive. use -D also.)
−
    -equil_error relerr iter: maximum relative equilibrium error allowed during the  
 
−
                    initial calculation of concentrations from the user specified
 
−
                    free ion concentrations and total buffer concentrations.
 
−
                    rel_error = ABS((true_kd - calculated_kd)/true_kd). Default = 1e-05
 
−
                    This should be small enough unless you have huge buffer
 
−
                    concentrations (>100 mM?). iter (an int) is maximum number of iterations
 
−
                    to allow during equilibration, max =1 billion, default = 10000000
 
  
−
Options controlling changing conditions:
+
  scale factors:
−
    -ion_src name current dur lowdisk highdisk lowring highring:  
+
-d Ryrimage1d.i2i #: the original intensity image for Ryr_image (since Ryr_image has pixel values as id numbers)
−
                    have a current source of the specified ion coming in through the specified rings
+
# is the number of Ryrimage1d.i2i intensity units required to produce spark current
−
                    and numdisk (rings and disks are 0-indexed indices, inclusive).
+
used in the calcium simulation (which produced the -f filename data).
−
                    Current is in pA for the specified duration (sec).
+
This will scale the [ca] produced by each Ryr voxel. Default = 30000.000000
−
                    The current is uniformly divided within the entire specified input volume.
+
-D BKimage2d.i2i #: the original intensity image for BK_image (since BK_image has pixel values as id numbers)
−
                    All -buffer and -ion options must be specified prior to this.
+
# is the BKimage2d.i2i intensity from one BK channel.
−
                    See comments by the -time_rate option.
+
This will scale current produced. Default = 1000.000000
−
    -ion_src_file name imagename current duration:
+
If you want the intensity of each Ryr or BK object to affect the ca release (for Ryr)
−
                    Similar to -ion_src, but the specified floating point image is used to
+
or number of channels (for BK) , you need to provide the original data by using the
−
                    scale the input current at each voxel. The total current coming into the
+
-d and -D options. Otherwise all voxels in an object contribute equally (so large objects
−
                    cell is adjusted so that after this per pixel rescaling, it will = "current".
+
have more receptors or channels than small ones).
−
                    -ion_current_trace with op = S, will modify this.
+
-c #: BK single channel conductance in pS, default = 220.0
−
                    Typically do: int2float -M 1 psf.i2i imagename        prior to this.
+
-C #: peak current (in pAmps) used in the simulation to produce -f filename, default = 1.000000
−
    -ion_current_trace name filename dt op threshold:  modifies current specified with
+
This is just used for output produced by -v to estimate spark current produced per Ryr cluster.
−
                    the -ion_src or -ion_src_file options.
 
−
                    Times which the channel is open (i.e., current flows) is specified in a file.
 
−
                    The file is a list of numbers (current or scale), 1 per line, at dt (in sec) intervals.
 
−
                    If op = T, currents greater than the specified threshold means channel open, otherwise closed.
 
−
                    If op = S, the current as specified with -ion_src is scaled by the number in the list.
 
−
                    "threshold" is required but ignored when op = S.  Not extensively tested.  
 
−
    -zap_sub name time:
 
−
                    At the specified time (in seconds) set the named substance concentration
 
−
                    to 0. This does not prevent it from being produced from other reactions.
 
−
                    This is useful if an injected "ion" was photons (e.g., photon+caged -> uncaged)
 
−
                    and the laser is turned off at the specified time.
 
  
−
Options related to output:
+
  miscellaneous:
−
    -save name filename type scale dt:
+
  -v: verbose. produces list of each Ryr cluster and K current produced to stderr. Other stuff too.
−
                    Save images of the specified substance (name) to specified file. By default
+
To write this to a separate file do: (channel_current -v ... > stdout.channel) >& stderr.channel
−
                    substances are saved in mM.  This will be multiplied by the scale
+
  -V: very verbose
−
                    specified here. dt (in sec) is how often to save images.  Type is I
 
−
                    (short int, our normal format) or F (floating pt,2x space, more accurate).
 
−
                    The first image saved will be prior to any ion current. The last image
 
−
                    will always be the end of the simulation. "name" should be as
 
−
                    specified with the -ion, -buffer or -kin option.
 
−
                    If the image file already exists, new images will postpended to it.
 
−
    -save all tag type scale dt:
 
−
                    Equivalent to -save for all ions and buffers. Uses default file names,
 
−
                    but adds the tag on the end. ions in nM, all else in uM, then times scale.
 
−
    -save_flow name rfilename lfilename scale dt:
 
−
                    Save flow images of the specified substance (name) to specified files.
 
−
                    rfile is radial flow, lfile is lengthwise flow. Images saved in floating pt.  
 
−
                    Flow is in pA (1 pA = 3 molecules/usec) times scale.
 
−
                    dt (in sec) is how often to save images. 
 
−
    -save_flow all ltag rtag scale dt:
 
−
                    Equivalent to -save_flow for all ions and buffers. Uses default file names,
 
−
                    but adds the tag on the end. Output same scale as above.
 
−
                    NOTE: flow for EACH substance takes 8*(number of voxels) of space.
 
−
    -debug #:      set debug level to #. May substantially slow down the simulation.
 
−
                    1: extra print statements at init. 2: more testing while running. 3:
 
−
    -debug_subvolume #: Normally the total mmoles (in the entire cell) of each saved substance is
 
−
                    printed to stderr. Setting this option will also cause the printing of the
 
−
                    total over a subvolume. Subvolume is all y >= # (0-indexed).
 
−
                    Example: if a cell has patch region at y=[1,10], setting # to the neck (11) of the
 
−
                    patch will let you see how much actually gets out into the main part of the cell.
 
  
−
Options related to MPI (multiprocessor stuff):
+
Examples:
−
    invoke on the wolves as, for example,  mpirun -np 3 -machinefile file_of_machine_names cellsim ...
 
−
    -print_proc #:  only print output from the specified process (0 indexed).
 
−
                  Some output (eg, Total mmoles of ions, etc) are only valid for process 0.
 
−
    -overlap #:  overlap neighboring regions by the specified number of voxels (default = 50)
 
−
                  This will also be the number of iterations between necessary synchronization.
 
−
    -debug_mpi #:  print MPI info. Execute MPI dummy stubs if MPI NOT present (eg, under Windows,SGI).
 
−
                  This version of the executable does NOT use MPI.
 
−
    -debug_mpi_proc rank numproc:  set dummy rank and number of processes. use only if MPI NOT present!
 
  
−
EXAMPLES
+
cellsim -spacing /home/lml/krypton/Zhuge/STIC/simulations/spacing2.txt ....     : produces ca.i2i
−
./cellsim -time .0010 -time_rate .5 -length 200 -rings 40 -radius 1 \
+
printvalsL -spacing2.txt -R -t -i ca.i2i > ca.vals    
−
        -ion Ca 100E-6 2.5E-6 2 -buffer Fluo3 .050 2.5E-7 -buffer Buf .230 1E-14 \
+
kate ca.vals     : delete fist comment line
−
        -kin Fluo3 Ca CaFl 8E4 90 2.5E-7 -kin Buf Ca CaBuf 1E5 100 1E-14        \
+
      objs_via_ascent -s 5 100 Ryr.i2i Ryrid.i2i > tmp
−
        -ion_src Ca 1 .00105 99 102 0 3 \
+
tail tmp  : the mean IOD gives an idea of brightness needed to produce current used in cellsim, say 30000
−
        -save all _run1 I 1 .0001 >& cellsim_run1.out
+
      objs_via_ascent -s 5 100 BK.i2i BKid.i2i > tmp
 +
tail tmp : the mean IOD gives an idea of brightness of a BK cluster, eg mean= 10000, but assume that's 10 channels
 +
 
 +
channel_current -f ca.vals -w 1 1 3 -d Ryr.i2i 30000 -D BK.i2i 1000 Ryrid.i2i Bkid.i2i > ryr_bk.info
 +
cull -s -c -1 -4 < ryr_bk.info |xmgrace -source stdin
 +
See also:
 +
closest_object closest_voxel objs_via_ascent /home/lml/krypton/Zhuge/BKandRyr/graph.pl,match.pl
 +
graph_channel_current.pl, test_channel_current, /home/lml/krypton/Zhuge/STIC/simulations/run32 
 +
/home/lml/krypton/Zhuge/BKandRyr/analysis6/cluster_distances.pl, plot_mean_sd_curve.pl
 +
/home/lml/krypton/Zhuge/xpp/ASM15.ode
 +
Bugs:
  
 +
Source code in ~krypton/lml/facil 
 +
# Popen is monotonic as it should be.
 +
must provide -f cafilename
  
−
Simulate for 100msec saving images every msec. 1 pA comes in the the pattern as specified
+
</nowiki>
−
by psf_float.i2i for 10 msec.  The current is then modulated by values in current.txt which
+
===  cluster ===
−
are one value per line, dt of 1 msec between them. Cell shape in mask.i2i. Voxel sizes in
+
  <nowiki>
−
spacing.txt
+
Drops random Ryr receptors into a world.  Then calculates how many clusters and their sizes get produced.
−
./cellsim -time .100 -time_rate .5 -spacing spacing.txt -mask mask.i2i \
+
Currently the "world" default size is  (2um)^3. (see -wsize)
−
        -ion Ca 100E-6 2.5E-6 2 -buffer Fluo3 .050 2.5E-7 -buffer Buf .230 1E-14  \
+
The origin of this world is in it's center (so it goes from -1um to +1 um in each direction by default).
−
        -kin Fluo3 Ca CaFl 8E4 90 2.5E-7 -kin Buf Ca CaBuf 1E5 100 1E-14        \
+
A histogram of cluster sizes is printed to stdout.
−
        -ion_src_file Ca psf_float.i2i 1 .010 \
 
−
        -ion_current_trace Ca current.txt .001 S 999 \
 
−
        -save CaFl cafl.i2i I 1000 .001 -save Ca ca.i2i I 1E6 .001 \
 
−
        -save_flow Ca ca_radial_float.i2i ca_long_float.i2i 1 .001 >& cellsim.out
 
−
 
 
−
SEE ALSO
 
−
      ~lml/vision/bin/float2int                    (convert a floating point image to standard SHORT INT format)
 
−
      ~lml/vision/bin/int2float       (convert normal image to floating point)
 
−
      ~lml/vision/bin/playexact                    (to display a floating point image)
 
−
      ~lml/invitro/Kargacin/changeheader          (change header of floating point image)
 
−
      ~lml/vision/bin/cyl2cart                    (convert from cylindrical to cartesian coords)
 
−
      ~lml/vision/bin/nadiffl7                    (many more options, but only 1 ion and 2 buffers)
 
−
      ~lml/vision/bin/diffuse2                    (simple Gaussian spot diffusion program)
 
−
      ~lml/invitro/Kargacin/iplot                  (3D graph of a 2D image, intensity -> height)
 
−
      ~lml/invitro/Kargacin/blur4D_dFoverF        (blur with psf, calculate dFoverF)
 
−
      ~lml/invitro/Kargacin/diffusion_limited_binding  (simulate diffusion limited binding)
 
−
      ~lml/invitro/Kargacin/spot_diffusion        (simulate diffusion from a spot)
 
−
      ~lml/vision/facil/printvals                  (print image values, calculate dFoverF)
 
−
      ~lml/vision/JSinger/src/channel              (steady state based upon Neher paper)
 
−
 
 
−
===  channel_current2 ===
 
−
channel_current2: like channel_current, but looks at [ca] over time at each BK (not just max [ca]).
 
−
From that, it calculates the max BK current. Channel_current adds the max [ca] that each BK voxel sees
 
−
from each voxel in a Ryr cluster, to get the max [ca] that BK pixel sees. But the max [ca] could occur
 
−
at different times since the Ryr cluster has a spatial extent.  Experimental.
 
−
 
 
−
Calculates the current which would be produced when each object in Ryr_image.i2i releases calcium which
 
−
activates nearby BK channels in BK_image.i2i. Produces a histogram of these output currents.
 
−
The input images should have each pixel in an object having the object id as its value. These images can be
 
−
produced with "countobjs -Q" or "objs_via_ascent"   
 
−
note:  a one pixel border around the image is ignored (added 12/8/09).
 
−
This may take about one-half hour to run.
 
  
 
Usage:
 
Usage:
−
channel_current [options] -f filename Ryr_image.i2i BK_image.i2i > im1_im2.hist
+
cluster [options]  
 
Options:
 
Options:
−
-f filename resting: file containing [ca], in nM, as a function of distance and time. Right now this is
+
distance options:
−
assumed to have 20 nm spacing and entries every msec. See
+
-eucliddist:     calculate a Euclidean distance (same in x,y,z directions). This is the default.
−
/home/lml/krypton/Zhuge/STIC/simulations/run32.ca as an example
+
  (use -cdist option to set the nearby distance for this).
−
"resting" is the resting [ca], in nM, for the cell. It will we subtracted from the other
+
-absdist:   rather than calculate a real distance, calculate max(ABS(dx),ABS(dy),ABS(dz)) - for speed
−
[ca] values to produce a "delta" [ca] which gets scaled linearly by Ryr intensity.
+
  (use -cdist to set the nearby distance for this.)
−
The spark current used to produce this (see -C) should be roughly in the range expected
+
-psfdist x y z:  distance (in um) a pt can be in the x, y, and z directions and be considered nearby.
−
for Ryr_image.i2i, since all delta [ca] is linearly extrapolated.
+
  Does not use -cdist.  To approximate a psf you might specify .2 .2 .6
−
All distances > largest distance in filename will have their [ca] set to the [ca]
+
      -cdist #:        how close (in um) two receptors must be to cluster, default = 0.500000
−
at the largest distance in filename.
+
other options:
−
note: Only the [ca] values at y == 1 are used. Is this what should be used?
+
      -2D:   do a 2D simulation (x and y), not a 3D simulation
−
-u #: voltage (mV). BK conductance is a function of voltage. default = -80.000000
+
-cylinder inner outer:  only place Ryr within a cylinder (e.g., plasma membrane) with specified
−
-U low delta hi: voltages from low to hi by delta. Max is 20 voltages
+
                        inner and outer radius (in um). The cylinder has its length the entire y dimension.
−
-r #:         resolution (binsize) for output histogram in femtoAmps, default = 1000.000000
+
  Only the parts of the cylinder which fit in the world (-wsize) are used.
−
-R #: reveral potential of BK channel in mV, default = -80.000000
+
-wsize x y z:   the width of the world (in um) in the x, y, and z directions (default = 2.0,2.0,2.0)
 +
-psize x y z:   pixel width (in um) in the x, y, and z directions (default = 0.1,0.1,0.1)
 +
      -numryr #:       number of Ryr receptors to drop, default = 1000
 +
-mindist #:   when placing Ryr receptors, keep this minimum distance (in um) between them.
 +
-randstart #:   start the random number generator with # as the seed. So runs can be reproduced.
 +
-image name.i2i   write out an image of the Ryr receptors.  Can then use blur3d to create a realistic
 +
  microscope image, and countobjs to analyze it and compare to real data.
 +
diagnostics:
 +
      -debug:          turn on debugging
 +
      -verbose:        be a bit more verbose
 +
      -h:  print this help
 +
      -H:  print this help
 +
      -help:  print this help
  
−
  distances:
+
Examples:
−
-w # # #: x,y, zweights for pixel distances (ie, pixel size), default = 1 for each.
 
−
-p #: pixel size in nm (i.e., what 1 means in -w option), default = 80.000000
 
−
-m # :         maxdistance (in units used for -w, i.e., pixels) to look for the closest pixel, default = 10.000000
 
  
−
  which pixels or clusters to include:
+
Source code in ~krypton/lml/Zhuge/Ryr_clusters 
−
-t # #: image1 and image2 thresholds, pixels > # are above threshold, default = 0.
+
  /home/lml/krypton/bin/cluster -h
−
-e lowx lowy lowz hix hiy hiz: only look at voxels within this extent (subvolume), 1-indexed, inclusive.
 
−
-s minsize maxsize:    only look at objects in Ryr_image.i2i within the size range (inclusive, in voxels)
 
−
max size allowed = 100000
 
−
-S minsize maxsize:    only look at objects in BK_image.i2i within the size range (inclusive, in voxels)
 
−
-i miniod maxiod:    only look at objects in Ryr_image.i2i within the iod range (inclusive. use -d also.)
 
−
-I miniod maxiod:    only look at objects in BK_image.i2i within the iod range (inclusive. use -D also.)
 
  
−
  scale factors:
+
</nowiki>
−
-d Ryrimage1d.i2i #: the original intensity image for Ryr_image (since Ryr_image has pixel values as id numbers)
+
=== diffusion_limited_binding  ===
−
# is the number of Ryrimage1d.i2i intensity units required to produce spark current
+
<nowiki>
−
used in the calcium simulation (which produced the -f filename data).
+
unknown option h
−
This will scale the [ca] produced by each Ryr voxel. Default = 30000.000000
 
−
-D BKimage2d.i2i #: the original intensity image for BK_image (since BK_image has pixel values as id numbers)
 
−
# is the BKimage2d.i2i intensity from one BK channel.
 
−
This will scale current produced. Default = 1000.000000
 
−
If you want the intensity of each Ryr or BK object to affect the ca release (for Ryr)
 
−
or number of channels (for BK) , you need to provide the original data by using the
 
−
-d and -D options. Otherwise all voxels in an object contribute equally (so large objects
 
−
have more receptors or channels than small ones).
 
−
-c #: BK single channel conductance in pS, default = 220.0
 
−
-C #: peak current (in pAmps) used in the simulation to produce -f filename, default = 1.000000
 
−
This is just used for output produced by -v to estimate spark current produced per Ryr cluster.
 
  
−
  miscellaneous:
+
Prints out distance (um) and concentration (particles/cm^3) at that distance
−
  -v: verbose. produces list of each Ryr cluster and K current produced to stderr. Other stuff too.
+
a slab volume of initial uniform concentration (1 particle/cm^3)which has a ligand
−
To write this to a separate file do: (channel_current -v ... > stdout.channel) >& stderr.channel
+
binding to a plane at x = 0. This is diffusion limited binding. See my notes of 10/20/98.
−
  -V: very verbose
 
  
−
Examples:
+
Usage: diffusion_limited_binding [options] 
 +
options:
 +
  -D #:  diffusion (cm^2/msec), default = 2.2E-09
 +
  -t t1 t2:  start and stop time to simulate (msec), default = 0.000000 1000.000000
 +
set both numbers equal if you only want one time point.
 +
  -x x1 x2: start and stop distances to print (um), default = 0.000000 10.000000
 +
set both number equal if you only want one position
 +
  -L #: thickness of slab volume (um), default = 10.000000
 +
  -T #: number of samples in t, default = 10
 +
  -X #: number of samples in x, default = 1000
 +
  -h: print this help message.
 +
  -F: print out time (msec) and flux (particles/cm^2-msec) at x = 0
 +
(i.e., rate molecules binding to x=0 surface).
 +
I've made flux towards lower x positive (normally it would be negative).
 +
  -B: print out time (msec) and total # of particles/cm^2 bound at x = 0
 +
(i.e., brightness of surface). Integral of flux from t1 = 0 regardless of -t option setting.
 +
  -s: don't print out concentration as a function of x (i.e, the entire slab)
 +
 +
</nowiki>
 +
===  ellipse  ===
 +
<nowiki>
  
−
cellsim -spacing /home/lml/krypton/Zhuge/STIC/simulations/spacing2.txt ....     : produces ca.i2i
+
This program creates an elliptical cell, cell length is in the y direction.
−
printvalsL -spacing2.txt -R -t -i ca.i2i > ca.vals    
+
It is used to model a squashed cell, and see how different it looks after
−
kate ca.vals     : delete fist comment line
+
blurring than a cylindrical cell
−
      objs_via_ascent -s 5 100 Ryr.i2i Ryrid.i2i > tmp
 
−
tail tmp  : the mean IOD gives an idea of brightness needed to produce current used in cellsim, say 30000
 
−
      objs_via_ascent -s 5 100 BK.i2i BKid.i2i > tmp
 
−
tail tmp  : the mean IOD gives an idea of brightness of a BK cluster, eg mean= 10000, but assume that's 10 channels
 
  
−
channel_current -f ca.vals -w 1 1 3 -d Ryr.i2i 30000 -D BK.i2i 1000 Ryrid.i2i Bkid.i2i > ryr_bk.info
+
Usage: ellipse [options] ellipse.i2i
−
cull -s -c -1 -4 < ryr_bk.info |xmgrace -source stdin
+
options:
−
See also:
+
  -d xdim ydim zdim:  dimensions of image, default = (50,100,50)
−
closest_object closest_voxel objs_via_ascent /home/lml/krypton/Zhuge/BKandRyr/graph.pl,match.pl
+
  -r xradius zradius:  radius (in pixels) in the x and z directions, default (20.000000,20.000000)
−
graph_channel_current.pl, test_channel_current, /home/lml/krypton/Zhuge/STIC/simulations/run32  
+
  -s samplerate:  number of subsamples in x and z directions per pixel, default = 1
−
/home/lml/krypton/Zhuge/BKandRyr/analysis6/cluster_distances.pl, plot_mean_sd_curve.pl
+
  -i intensity:  intensity inside the ellipse, default = 100.000000
−
/home/lml/krypton/Zhuge/xpp/ASM15.ode
+
−
Bugs:
+
</nowiki>
 +
===  em  ===
 +
  <nowiki>
 +
Creates (or reads in) a HMM and then generates random data from it
 +
(each model is a poisson process, with the j_th model having a lambda = 10*j).
 +
Then attempts to use an EM algorithm to estimate the original model.
 +
This is a way for me to learn how to implement the EM algorithm.
  
−
Source code in ~krypton/lml/facil  
+
Usage: em [options]
−
  # Popen is monotonic as it should be.
+
options:
−
must provide -f cafilename
+
  -s #: number of states, default = 2. Must specify -t and -p after this.
−
 
+
  -t # # # ... #: state transition probabilities. If there are S states (e.g., -s #)
−
=== cluster ===
+
                        order: T[1][1], T[1][2], ... T[1][S], T[2][1] ... T[S][S] if there are S states),
−
Drops random Ryr receptors into a world.  Then calculates how many clusters and their sizes get produced.
+
                        where T[i][j] is the probability of a transition from state i to state j.
−
Currently the "world" default size is (2um)^3.  (see -wsize)
+
  -p # # # ... #: prior probabilities of states (S entries if there are S states)
−
The origin of this world is in it's center (so it goes from -1um to +1 um in each direction by default).
+
  -P #: which prior estimate to use for the pdf of the data (ie., estimate of Poisson process)
−
A histogram of cluster sizes is printed to stdout.
+
        1 = uniform, 2 = delta, 3 = uniform+delta, 4 = peice of observed data histogram (default)
 +
        5 = true pdf (Poisson)
 +
  -n #: number of data points to simulate, default = 100
 +
  -m #: maximum number of iterations to perform, default = 20
 +
        -truetrans: use the true transition probabilities as the initial guess (default = uniform)
 +
        -trueprior: use the true prior probabilities as the initial prior guess (default = uniform)
 +
  -findstates:  also find (and print) the estimate of the states at each time point.
 +
  -v:  verbose. print more info to stderr.
 +
  -V:  print some more info. Some printouts only occur if # data pts is < 1000
 +
 
 +
Source code in /home/lml/krypton/facil/em.cc
 +
See also:  ~/krypton/facil/transpose.pl  to print out selected lines from the output file
 +
          ~/krypton/matlab/netlab: has EM routines
 +
          ~/krypton/matlab/bayes/BNT/HMM: has HMM and EM routines
 +
/home/lml/krypton/bin/em -h
 +
 
 +
</nowiki>
 +
===  makebeads2  ===
 +
<nowiki>
 +
/home/lml/krypton/bin/makebeads2
 +
 
 +
randomly distributes beads within a volume.  created to let Jack simulate
 +
distribution of glut4 labels vesicles in the TIRF field of a microscope.
 +
A bead has a constant value between its inner and outer diameters. This value
 +
falls to zero 3 standard deviations inside the inner diameter and 3 sd outside
 +
the outer diameter. Beads are not allowed to intersect each other.
 +
Writes info about beads created or moved to stdout (can be read back in with -b option).
 +
note: coords written out are zero indexed (not 1-indexed like addlines and play) in pixels.
 +
usage:
 +
makebeads2 [options] outimage.i2i
 +
options:
 +
-d x y z: image dimensions in pixels, default = (256,256,64)
 +
-D # #: bead inner and outer diameters in microns, default = 0.200000 and 0.200000
 +
  -R #: inner rise standard deviation (in microns), default = 0.012500
 +
  -F #: outer fall standard deviation (in microns), default = 0.012500
 +
-S x y z: x, y, and z pixel dimensions (in microns), default = (0.040000,0.040000,0.010000)
 +
-c x y z: x, y, and z pixel dimensions for new image size.  If you are planning on taking the
 +
image produced by makebeads2 and using chres to change pixel size, put the new
 +
pixel sizes here. This is used when calculating the intensity of the brightest pixel
 +
from each bead. This output info can be useful for signal to noise ratio tests.
 +
-X #: number of subpixels per pixels, default = 10
 +
-p # val: set z plane # (0 indexed) to val (modelling diffuse fluorescence on the plasma membrane)
 +
If val is < 1 it is interpreted as a fraction of total light in the image.
 +
For example: val = .2 means set the z plane to the intensity necessary so that the total
 +
light in that plane is 2012f the total in the final image (i.e., after exponential if -e or -E)
 +
-e dist: apply an exponential scale factor exp(-z/dist) to each zslice; like a TIRF excitation intensity
 +
dist is the 1/e distance in microns (.1 is a good value)
 +
This exponential also get applied to z plane value set via "-p # val" if val >= 1. It does
 +
NOT apply to that if val < 1.
 +
  -E amp2 dist2:apply a second an exponential scale factor amp2*exp(-z/dist2) to each zslice
 +
must also use -e option. Each intensity will be scaled by (amp1*exp(-z/dist)+amp2*exp(-z/dist2)).
 +
amp1 + amp2 = 1 and is automatically set when -E is used (amp1 = 1-amp2).
 +
 
 +
options related to output:
 +
-i #: outimage.i2i should be scaled by # before writing it.  Format is short int (our default format).
 +
-f #: outimage.i2i should be scaled by # before writing it.  Format is float.
 +
-v: verbose.
 +
-A: don't create outimage.i2i (still need a dummy name on command line though).
 +
-B: don't even simulate voxels (just prints beads centers, amplitudes, sizes).  Automatically sets -A.
 +
-u file.objs: create file, which has a list of bead objects created, in the same format countobjs produces.
 +
this lets utilities written to analyze files in that format to then be used
 +
(e.g., objs2dist.pl, objs2bb.pl). COM field is the bead center even if part of it is outside
 +
the image. Size field is in pixels (only counting what is within the image).
 +
IOD is the total bead light in the image after exponential weighting (if -e or -E) and scaling (-i, -f).
 +
AVG = IOD/size.  MAX is the max pixel (after -e,-E,-i,-f) . Some file header info may be wrong.
 +
-U file.objs: like -u, but all z coords are replaced by 1, and coordinates are for the new resolution specified
 +
by -c option. MAX value and max coord come from a sum projection in z at the new resolution.
 +
(after -e,-E,-i,-f).
 +
-W: print some warning messages to stderr if appropriate.  note: This will mess up the output in the file
 +
if you are running it as:  makebeads2 -W .... >& file.out
 +
 
 +
options controlling randomization (of a uniform random distribution unless otherwise noted):
 +
-s min max: mininum and maximum range of allowed bead sizes. This number scales the
 +
bead inner and outer diameters (does not change sd). default = 1.000000 and 1.000000
 +
-G mean sd:  make bead sizes (scales) a Gaussian variable with specified mean and sd (instead of -s).
 +
note: since beads can't intersect, if you place beads densely you will tend to get more
 +
small beads (since the program generates a new random bead if one doesn't fit), and hence
 +
the size distribution will no longer match the parameters specified here.
 +
-g mean sd shell: like -G, but also specify bead shell thickness in microns.  mean doesn't scale the shell thickness.
 +
mean is the diameter to midway between the inner and outer diameters (so don't use -D option).
 +
-a min max: min and max amplitude (brightness) of bead, default = 1000.000000 and 1000.000000
 +
a pixel which is entirely covered by the shell of the bead (between -D numbers) will have this value.
 +
So this is like specifying a fluorescent concentration.
 +
-P lambda scale: make amplitude a Poisson variable with the specified mean (lambda) value (instead of -a option).
 +
this value then gets multiplied by scale.  Amplitudes of 0 get ignored (a new amplitude is calculated).
 +
note: this is still just the amplitude of a covered pixel, not of the entire bead unless -T used too.
 +
-T: The amplitude specified with -a or -P option is the total light in the bead, not the light
 +
of one fully covered pixel.  Takes about double the time to run.
 +
note: if the amplitude isn't large compared to number of pixels in the bead's shell, then
 +
individual pixel intensities will be small and truncation may effect total intensity.
 +
-x min max:  allowed position of beads, in pixels, zero indexed, inclusive. The entire bead must fit in this region
 +
unless -o option also specified.
 +
-y min max:  allowed position of beads, in pixels, zero indexed (default = anywhere within image)
 +
-z min max:  allowed position of beads, in pixels, zero indexed (default = anywhere within image)
 +
-o: allow bead centers to fall outside the image volume (except in low z direction, i.e. coverslip).
 +
This is a more accurate way to represent a small view of a large cell.
 +
This option is ignored if -b option used.
 +
-O: like -o, but beads in low z direction outside the volume reflect back inside the volume (i.e. get
 +
moved back inside volume instead of thrown away).
 +
note: if bead density is too high, reflecting a bead inside may cause it to intersect with a previous
 +
bead, in which case the reflecting bead will be thrown away and a new randomly placed bead created.
 +
-w #: use # as the seed for random number generator, so can reproduce exact runs. must be negative.
 +
-b file xyspeed zspeed:  read initial bead params (e.g., size, position) from file, just add random position to this.
 +
the random position uses the speeds (in microns) as the max the vesicle can move in each direction
 +
in this time period (i.e., dx = ran(0 to 1)*xyspeed, etc).
 +
Set xyspeed and zspeed = 0 if you don't want to move the beads.
 +
  -j #: long int starting seed for random number generator. specify the same number (not 0) each time you run this
 +
program if you want to exactly repeat the random numbers generated. The default is to use a different
 +
starting seed each time based upon the process id when you run the program.
 +
-n #: number of beads to make, default = 100, max = 10000
  
−
Usage:
+
Examples:
−
cluster [options]
+
#create 200 beads, with a 50 nm (5 z plane) gap before the pm, which has an
−
Options:
+
#intensity of 50. Apply a TIRF exponential excitation scale to it.
−
distance options:
+
makebeads2 -e .1 -z 5 63 -p 5 50 -n 200 beads1.i2i > beads1.info
−
-eucliddist:      calculate a Euclidean distance (same in x,y,z directions). This is the default.
+
# apply microscope psf
−
  (use -cdist option to set the nearby distance for this).
+
blur3d -P -N 256 256 128 beads1.i2i empiricalpsf_seg_40nm_seg.i2i beads1_blur3d.i2i
−
-absdist:   rather than calculate a real distance, calculate max(ABS(dx),ABS(dy),ABS(dz)) - for speed
+
# bin (averge) to 100/160 nm resolution
−
  (use -cdist to set the nearby distance for this.)
+
reduceima -A -X 1 256 4 -Y 1 256 4 beads1_blur3d.i2i beads1_blur3d4x4.i2i
−
-psfdist x y z:  distance (in um) a pt can be in the x, y, and z directions and be considered nearby.
+
# extract infocus plane
−
  Does not use -cdist. To approximate a psf you might specify .2 .2 .6
+
segment -Z 16 16 1 beads1_blur3d4x4.i2i beads1_blur2d4x4.i2i
−
      -cdist #:        how close (in um) two receptors must be to cluster, default = 0.500000
+
# add ScratchNG camera noise
−
other options:
+
noise -e 1.4 -n 8 beads1_blur2d4x4.i2i beads1_blur2d4x4N.i2i
−
      -2D:   do a 2D simulation (x and y), not a 3D simulation
+
histima -size 10 -bins 100 beads1_blur2d4x4N.i2i > beads1_blur2d4x4N.hist
−
-cylinder inner outer:  only place Ryr within a cylinder (e.g., plasma membrane) with specified
+
 
−
                        inner and outer radius (in um). The cylinder has its length the entire y dimension.
+
#creates beads, move them, look at the results
−
  Only the parts of the cylinder which fit in the world (-wsize) are used.
+
makebeads2 -z 5 63 beads1.i2i > beads1.info
−
-wsize x y z:   the width of the world (in um) in the x, y, and z directions (default = 2.0,2.0,2.0)
+
#read those beads in and move them. but still must stay outside the first 50 nm.
−
-psize x y z:   pixel width (in um) in the x, y, and z directions (default = 0.1,0.1,0.1)
+
makebeads2 -z 5 63 -b beads1.info .05 .05 beads2.i2i > beads2.info
−
      -numryr #:        number of Ryr receptors to drop, default = 1000
+
concateima beads1.i2i beads2.i2i beads.i2i
−
-mindist #:   when placing Ryr receptors, keep this minimum distance (in um) between them.
+
imsets beads.i2i 2
−
-randstart #:   start the random number generator with # as the seed. So runs can be reproduced.
+
dave -tdim 2 -4 -I beads.i2i
−
-image name.i2i   write out an image of the Ryr receptors.  Can then use blur3d to create a realistic
+
 
−
  microscope image, and countobjs to analyze it and compare to real data.
+
#just create scripts (very fast) with positions of beads, and move them around.
−
diagnostics:
+
makebeads -z 5 63 -n 200 -A dummy.i2i > beads1.info
−
      -debug:          turn on debugging
+
makebeads -b beads1.info .05 .05 -z 5 63 -A dummy.i2i > beads2.info
−
      -verbose:        be a bit more verbose
+
makebeads -b beads2.info .05 .05 -z 5 63 -A dummy.i2i > beads3.info
−
      -h:  print this help
+
#now create the images
−
      -H:  print this help
+
makebeads -b beads1.info 0 0  beads1.i2i > beads1.info2
−
      -help:  print this help
+
makebeads -b beads2.info 0 0  beads2.i2i > beads2.info2
 +
makebeads -b beads3.info 0 0  beads3.i2i > beads3.info2
 +
 
 +
see also: makebead, readbeads.pl, blur3d, project, /storage/big1/sh/simul
 +
 
 +
source code in ~lml/krypton/facil
 +
 +
</nowiki>
 +
===  oxygen  ===
 +
<nowiki>
 +
Calculates steady state partial oxygen pressure (P) as a function of distance from a blood vessel (r).
 +
The blood vessel has a radius of R1 (5.0 um) and Oxygen at the vessel is Ps (52.0 mm Hg).
 +
The default is to print partial Oxygen pressure (mm Hg) as a function of distance (um) as columns to stdout.
 +
All models set P at R1 (and r<R1) to Ps. All models clamp P at R1 to be Ps for all time (vessels stay oxygenated).
 +
 
 +
The line source model (-m 1) implements equation 17 from KroghOxygenDiffusionModel1.pdf
 +
  Modeling pO2 Distributions in the Bone Marrow Hematopoietic Compartment.
 +
  I. Krogh's Model  by D.C.Chow, L. A. Wenning, W. M. Miller, and E.T. Papoutsakis
 +
  Biophysical Journal, vol. 81, August 2001, pp. 675-684.
 +
This models O2 assuming a constant oxygen consumption by the tissue and constant O2 (Ps) at the blood vessel.
 +
They claim results don't change by more than a few percent using proportional (or other) O2 consumption models.
 +
The value of dP/dr at R2 is forced to 0.
 +
note: this is an implmentation of O2 from a line source (e.g., blood vessel), not a point source.
 +
So it is not appropriate to convolve (in 3D) this with a 3D image of blood vessels.
 +
But -p (or -m 2) will produce a point source image.
 +
 
 +
The point source model (-m 2) implements equation 14 from HeatConduction.pdf
 +
  Steady Heat Conduction in Layered Mediums: The Half-Space and Sphere, by Henry N. Pollack
 +
  J. of Geophysical Research, vol. 70(22), Nov. 15, 1965, pp. 5645-5648
 +
This models O2 assuming a constant oxygen consumption by the tissue and constant O2 (Ps) at the blood vessel.
 +
note: The value of dP/dr at R2 is forced to 0 unless -b # is specified.
 +
note: Since O2 consumption is constant, it might not make sense to use this in a convolution, since it isn't
 +
      linear.
 +
 
 +
Model 3 is a point source model, but O2 consumption by the tissue is proportional to the O2 level in the tissue.
 +
  I derived it from Crank's equations 6.60 (infinite hollow sphere model) and 14.13 (see below).
 +
   It requires the specification of an initial O2 level in the tissue (P0, which is the same everywhere).
 +
  R2 is used only to determine how far out to plot the graph (and dP/dr at R2 is not used either).
 +
  See my paper notes or ~/krypton/Corvera/Olga/SteadyStatePointDiffusion.pdf
 +
  Not only is proportional O2 consumption more biological, but I think this should be linear and therefore
 +
  it makes more sense to use in a convolution with a (binarized?) blood vessel image.
 +
 
 +
note: because R2 really should vary in an image, depending upon the distance between nearby blood vessels
 +
        and because Ps is not just a scale factor (so, eg, brighter blood vessels in an image, if brightness is
 +
        proportional to Ps, do NOT just cause a scale factor in the P(r) values), it may NOT be appropriate
 +
to take the image produced by this program and convolve it with an image of blood vessels to get
 +
estimates of oxygenation of tissue. This comment may apply to all three models (definitely to -m 1)
 +
 
 +
Usage: oxygen -m # [options] 
 +
    required:
 +
  -m # which model to use.
 +
        -m 1 :line source model with constant O2 consumption
 +
-m 2: point source model with constant O2 consumption.
 +
        -m 3 :point source model with O2 consumption proportional to O2 concentration.
 +
Use point source models if planning on convolving resulting image with a blood vessel image.
 +
 
 +
    options for all models:
 +
Although these options can be used in all models, different models have different sensitivities to them.
 +
  -R1 # vessel radius (um). Changes to this can change P a lot (if -m 1), and may require R2 to change (see -c).
 +
default = 5.00
 +
  -R2 # max distance any tissue is from a vessel (um). This changes the solution a lot for some models
 +
but for the -m 3 model it only affects how far out data is graphed, default = 2268.00
 +
More precisely, for -m 1 and -m 2 it is the distance at which dP/dr = 0 (ie, a b.c. on the pde).
 +
  -Ps # partial pressure of Oxygen at the blood vessel (mm Hg). Default = 52.00
 +
40 is typical of veins, 95 of arteries. This is held constant over time in all the models right now.
 +
For -m 1, this will affect the O2 value at R1, but how P changes RELATIVE to P(at R1) (as r increases)
 +
will not be affected (ie, it does NOT just scale everything).
 +
    model 1 options:
 +
  -c use with -R1 to automatically calculate R2 based on the specified R1 (since R2 affects the solution).
 +
(this also sets -m 1 as a convenience)
 +
    model 2 options:
 +
  -b # changes the 2nd boundary condition to be P at R2 = # mm Hg
 +
instead of dP/dr = 0 at R2 (this also sets -m 2 as a convenience)
 +
    model 3 options:
 +
  -P0 # the initial oxygenation level of the tissue (mm Hg). Default = 0.000000
 +
This will change the solution a lot, since it is also the level that O2, in steady state, approaches as r->infinity.
 +
Therefore, nonzero P0 may also make convolution with a blood vessel image inappropriate.
 +
-P0 0 seems to produce the same results as -r. (-P0 also sets -m 3 as a convenience)
 +
 
 +
    output graph options:
 +
  -n normalize output (doesn't affect -i image.i2i), ie, print r/R1  vs P/Ps  rather than r vs. P
 +
  -r add a third reference column which is the "simple" diffusion solution (goes as 1/r)
 +
This is for a point source in 3D (not a line source), and ignores O2 consumption.
 +
graph as: oxygen_krogh -r | xmgr -nxy stdin
 +
 
 +
    output image options:
 +
  -i image.i2i xdim ydim zdim:  create an image of the values, to use with blur3d to find
 +
O2 everywhere in an image as a function of distance from blood vessels.
 +
If you are planning on convolving you should probably specify -m 3 .
 +
  -d x y z if -i is specified. pixel dimensions in um. default = 0.60 0.60 10.00
 +
  -s # if -i is specified. scale output intensities by #. Otherwise max is 52.0 mm Hg
 +
  -o xorig yorig zorig: location of the blood vessel in the image (zero indexed, floating point)
 +
blur3d may care about this, depending upon options. default = (0.00,0.00,0.00)
 +
(maybe I should change this to define a line in the image when -m 1 is specified?)
 +
-p the output image should be for the simple 1/r point source (see -r explanation) rather
 +
for the line source (-m 1) or point source with O2 consumption (-m 2 or -m 3).
 +
 
 +
    other options:
 +
  -v verbose. stuff to stderr.
 +
  -V more verbose. stuff to stderr. debugging info.
 +
  -h print this help
  
 
Examples:
 
Examples:
 +
      # create a binary image of blood vessels (maybe use BestPath3D_ellipse instead? Maybe thin3D afterwards?):
 +
    mask_image -m 2700 -r 1 G03_x_1_2_r4.i2i G03_x_1_2_r4.i2i G03_x_1_2_r4_binaryvessels.i2i
 +
      # create a psf of blood oxygenation:
 +
    oxygen -s 500 -i O2ptsource.i2i 50 50 21 -d 2.58 2.58 10 -o 25 25 10 -m 3 -r > O2ptsource.xy
 +
    blur3d -S .002 -Z -d -v G03_x_1_2_r4_binaryvessels.i2i O2ptsource.i2i G03_x_1_2_r4_O2.i2i
 +
 +
see also:  /home/lml/krypton/Corvera/Olga/README, SteadyStatePointDiffusion.pdf (my notes)
 +
        Random Walks in Biology (New, expanded edition) by Berg p. 23 eq. 2.11;
 +
The Mathematics of Diffusion (2nd Ed) by Crank p. 32 eq  3.5b; p. 89 eq 6.5; p. 102 eq 6.60; p. 330 eq 14.13
 +
source code in:  /home/lml/krypton/facil
 +
error: A model must be specified, i.e., -m # must be specified.
 +
  
−
Source code in ~krypton/lml/Zhuge/Ryr_clusters  
+
</nowiki>
−
  /home/lml/krypton/bin/cluster -h
+
===  sparkspread ===
 +
  <nowiki>
  
−
===  diffusion_limited_binding  ===
+
Calculates the spread of spark event amplitudes given that two events occur at the same
−
unknown option h
+
spark site and compares it to the spread of amplitudes given that two events occur at
 +
different spark sites. A spark site has a unique (x,y) position.
 +
Prints mean and stddev to stdout. Also prints out histograms and cumulative distributions.
  
−
Prints out distance (um) and concentration (particles/cm^3) at that distance
+
A valid datafile has either blank lines, comments (start with #) or a data line (spark event).
−
a slab volume of initial uniform concentration (1 particle/cm^3)which has a ligand
+
A data line has 8 or 11 fields. 11 if a stoc is associated with the spark event
−
binding to a plane at x = 0.  This is diffusion limited binding. See my notes of 10/20/98.
+
A valid stoc (last 3 fields) must have a nonzero stoc amplitude (1st of 3 fields)
 +
See /usr/people/lml/vision/Rhonghua/Sparks/singlesitev2 for a valid file format.
  
−
Usage: diffusion_limited_binding [options]
+
Usage: sparkspread [options] datafile
 
options:
 
options:
−
  -D #:   diffusion (cm^2/msec), default = 2.2E-09
+
  -h: do NOT print histogram for intraspark data.
−
  -t t1 t2:   start and stop time to simulate (msec), default = 0.000000 1000.000000
+
  -H: do NOT print histogram for interspark data.
−
set both numbers equal if you only want one time point.
+
  -c: do NOT print cumulative distribution for intraspark data.
−
  -x x1 x2: start and stop distances to print (um), default = 0.000000 10.000000
+
  -C: do NOT print cumulative distribution for interspark data.
−
set both number equal if you only want one position
+
  -b #: number of bins for histogram, default = 20
−
  -L #: thickness of slab volume (um), default = 10.000000
+
  -n: normalize the histograms so their maximum is 1
−
  -T #: number of samples in t, default = 10
+
  -k: also print ks statistic showing if two distributions are different.
−
  -X #: number of samples in x, default = 1000
+
−
  -h: print this help message.
+
</nowiki>
−
  -F: print out time (msec) and flux (particles/cm^2-msec) at x = 0
+
===  spot_diffusion ===
−
(i.e., rate molecules binding to x=0 surface).
+
<nowiki>
−
I've made flux towards lower x positive (normally it would be negative).
+
unknown option h
−
  -B: print out time (msec) and total # of particles/cm^2 bound at x = 0
 
−
(i.e., brightness of surface). Integral of flux from t1 = 0 regardless of -t option setting.
 
−
  -s: don't print out concentration as a function of x (i.e, the entire slab)
 
−
 
−
===  ellipse ===
 
  
−
This program creates an elliptical cell, cell length is in the y direction.
+
Prints out time (secs) and concentration (nMolar) seen at a distance
−
It is used to model a squashed cell, and see how different it looks after
+
from a channel with a constant current, open for specified time.
−
blurring than a cylindrical cell
 
  
−
Usage: ellipse [options] ellipse.i2i
+
Usage: spot_diffusion [options]
 
options:
 
options:
−
  -d xdim ydim zdim:  dimensions of image, default = (50,100,50)
+
  -D #:  diffusion (cm^2/sec), default = 2.2E-06
−
  -r xradius zradius: radius (in pixels) in the x and z directions, default (20.000000,20.000000)
+
  -c #:  current (pAmp), default = 1
−
  -s samplerate: number of subsamples in x and z directions per pixel, default = 1
+
  -s #:  starting concentration (resting level, in nM), default = 0
−
  -i intensity: intensity inside the ellipse, default = 100.000000
+
  -T #:   total time to simulate (seconds), default = 1.000000
 +
  -t #: length of time current is on (seconds), default = 1.000000
 +
  -r #: distance from channel opening (um), default = 0.100000
 +
  -R r0 r1 dr: average over a range of distances (crude psf), from r0 to r1 by dr (in um)
 +
  -n #: number of time points to sample, default = 1000.000000
 
   
 
   
−
===  em ===
+
</nowiki>
−
Creates (or reads in) a HMM and then generates random data from it
+
===  total_ca ===
−
(each model is a poisson process, with the j_th model having a lambda = 10*j).
+
<nowiki>
−
Then attempts to use an EM algorithm to estimate the original model.
+
#/home/lml/krypton/bin/total_ca
−
This is a way for me to learn how to implement the EM algorithm.
+
 
 +
Reads in a floating point image or short int image as produced by simulation (i.e., cylindrical coords), calculates
 +
the total calcium change (i.e., [ca](r)*pi*r*r - t(0)) as a function of distance from the calcium entry site.
 +
Output is printed to stdout.
 +
All input concentrations should be in nM
 +
Each z slice is analyzed separately (i.e., z is assumed to be time) unless image is rectangular and 3D (-R).
  
−
Usage: em [options]  
+
Usage: total_ca [options] ca.i2i
−
options:
+
      options for printing graphs of calcium (not concentration) vs. distance from ca entry location:
−
  -s #: number of states, default = 2. Must specify -t and -p after this.
+
the units for the graph are nM*um^3, but 1nM = .6ions/um^3 , so multiply be .6 to get ions
−
  -t # # # ... #: state transition probabilities. If there are S states (e.g., -s #)
+
                      (and 1pA = 3,000 ions/msec if you want to convert to pA-msec).
−
                        order: T[1][1], T[1][2], ... T[1][S], T[2][1] ... T[S][S] if there are S states),
+
        -n: normalize calcium distance graph for each z slice (so sum of amplitudes = 1),
−
                        where T[i][j] is the probability of a transition from state i to state j.
+
                      (otherwise when graph spacing is changed, -g option, the height of the graph will change).
−
  -p # # # ... #: prior probabilities of states (S entries if there are S states)
+
  -a:   print absolute calcium, not delta calcium (the default).
−
  -P #: which prior estimate to use for the pdf of the data (ie., estimate of Poisson process)
+
                      when this is applied to a uniform image it should produce a 4pi*r*r distribution.
−
        1 = uniform, 2 = delta, 3 = uniform+delta, 4 = peice of observed data histogram (default)
+
            The default, delta calcium, prints the delta (i.e., change) from slice 0 (regardless -z option)
−
        5 = true pdf (Poisson)
+
        (but delta calcium not calculated if 3D rectangular, i.e., -R specified).
−
  -n #: number of data points to simulate, default = 100
 
−
  -m #: maximum number of iterations to perform, default = 20
 
−
        -truetrans: use the true transition probabilities as the initial guess (default = uniform)
 
−
        -trueprior: use the true prior probabilities as the initial prior guess (default = uniform)
 
−
  -findstates:  also find (and print) the estimate of the states at each time point.
 
−
  -v:  verbose. print more info to stderr.
 
−
  -V:  print some more info. Some printouts only occur if # data pts is < 1000
 
  
−
Source code in /home/lml/krypton/facil/em.cc
+
  -p # #: pixel size in microns in the radial (x) and lengthwise (y) directions,
−
See also: ~/krypton/facil/transpose.pl  to print out selected lines from the output file
+
default =  0.10 and  0.10
−
          ~/krypton/matlab/netlab: has EM routines
+
  -P # # #: pixel size in microns in x,y,z directions (rectangular not cylindrical coords),
−
          ~/krypton/matlab/bayes/BNT/HMM: has HMM and EM routines
+
default = ( 0.10, 0.10, 0.10)
−
/home/lml/krypton/bin/em -h
+
  -r: data is rectangular, not cylindrical coords (-P sets this too)
 +
(e.g., already passed through cyl2cart).
 +
  -R: data is 3D rectangular (not 2D+time), not cylindrical coords.
 +
  -e x y:      0-indexed coords of calcium entry location (default: x= 0, y = ydim/2)
 +
  -E x y x:    0-indexed coords of calcium entry location for 3D rectangular data
 +
(i.e., z direction is not time). sets -R option.
 +
        -g #: graph spacing. default is the pixel size.
 +
        -s #:         subsample each voxel this number of times in the x (radial) and y (longitudinal) directions,
 +
                      (and z if 3D and not 2D+time) default = 3 (anything less tends to introduce errors).
 +
        -z low hi dz: only look at z slices low to hi (0-indexed, inclusive), stepping by dz.
  
−
== Statistics ==
+
see also:
−
===  cluster_distances.pl  ===
+
    printvals, float2int, in matlab: readxmgr, acurves (see ~lml/krypton/JSinger for an example)
  
−
  Takes output from channel_current2 (after 11/24/09) and pulls out statistics about
+
source code in ~lml/krypton/Walsh directory.
−
  mean distance to each BK cluster. Then produces average stats from all the files
+
   
−
  analyzed. Output to stdout.
+
</nowiki>
 +
=== vcspat-models  ===
 +
  <nowiki>
 +
  argc = 1
  
 +
This program creates a volume image with objects randomly placed
 +
within it.  The mask image defines where an object is allowed to be placed.
 +
The mask value can be changed with the -M option.  Objects can either be individual
 +
voxels (the default) or tiny images (-o option) or connected voxels in an image (-O).
  
−
usage:
+
Usage:
 +
  vcspat-models [options] mask.i2i out.i2i
  
−
   -files:file1:file2:...:filen       which files to analyze. Defaults are files named 2_*.currents in cwd.
+
Options:
−
   -numBKvsProb       also prints numBK vs prob data suitable for xmgrace
+
   -m #    : model number (default 0).
 +
            0 - grid pattern
 +
            1 - random pattern
 +
  -M #    : value of mask pixels (default 1).
 +
  -r #    : number of random points to use in random pattern (default 25000)
 +
  -o name  : image to use at point in pattern (default just turns on a pixel)
 +
(can use up to 256 images - image will be randomly chosen)
 +
   -a : if objects overlap, add their pixel values instead of just replacing
  
 +
Options related to objects:
 +
  -O thresh name.i2i: threshold an image to use for objects. Extracts as an object each
 +
            connected set of voxels above threshold.  Places these  objects randomly
 +
            into the output image (uses each object once). Use with the
 +
            -m 1 option. Do not use the -r option
 +
  -P name.i2i id.i2i: similar to -O option, extracts as an object each of the
 +
    the objects in name.i2i based upon their pixel values (object id) in id.i2i
 +
(e.g., id.i2i was produced via objs_via_ascent). This option preserves the
 +
distinction between objects in name.i2i which may be touching - since simple thresholding is not used.
 +
If only an id.i2i is available (no name.i2i), you can specify -P id.i2i id.i2i
 +
  -v: replace objects (after fitting them into the image) with just the single voxel
 +
at the brightest location in the object.  Only use with -O and -e options.
 +
  -S # #:  minimum and maximum object sizes to allow (use with -O option).
 +
  -e: do not allow (exclude) objects to be placed so that they overlap with each
 +
other, go outside the image, or go outside the mask.  The default is to allow
 +
overlaps and to just zero out that part of an object which goes outside the
 +
image volume or mask.
 +
  -n: number the objects in out.i2i (use with -O or -P options). Rather than all
 +
the pixels in an object being their original values, all the pixels in object 1
 +
will be labeled 1, all those in object 2 labeled 2, etc.  If -e not specied and
 +
objects overlap the first object placed (usually the largest) will prevail.
 +
Can also be used with -v, then output will have the brightest voxel set to object id, others to 0.
 +
The output of this can be analyzed by object_overlap program.
 +
  -N out2.i2i:  create a second output file, number the objects in out2.i2i (like -n).
  
−
example:
+
OtherOptions
−
   (channel_current2  .... > file.channel_current) >& file1.currents
+
  -s #    : seed for random number generator (default pid)
−
   (channel_current2  .... > file2.channel_current) >& file2.currents
+
   -x #    : x increment for grid pattern (default 3)
−
   cluster_distances.pl -numBKvsProb -files=file1.currents:file2.currents > avg_distances.txt
+
   -y #    : y increment for grid pattern (default 3)
−
   fgrep -v clusters avg_distances.txt |xmgrace -pipe
+
   -z #    : z increment for grid pattern (default 3)
 +
   -V: verbose
  
−
  note: mean distances to clusters are not always increasing (e.g., mean distance to 10th
+
note: entire image name (with extension) is required.
−
      cluster could be SMALLER than the mean distance to the 9th cluster).  I think this
+
note: a - in place of an image name means stdin or stdout.
−
      has to do with limiting my maximum distance (see comments in this file).
+
Examples:
 +
  objs_via_ascent in1.i2i inid1.i2i
 +
  objs_via_ascent in2.i2i inid2.i2i
 +
  vcspat-models-new -m 1 -M 1 -P in1.i2i inid1.i2i -N outid1.i2i mask.i2i out1.i2i
 +
  vcspat-models-new -m 1 -M 1 -P in2.i2i inid2.i2i -N outid2.i2i  mask.i2i out2.i2i
 +
  object_overlap outid1.i2i outid2.i2i  > obj_overlap
 +
  overlapL out1.i2i out2.i2i > coloc
 +
 
 +
See also: krypton/Qiong/run1,  vcspat, randompuncs, spatial_stats2, objs_via_ascent, countobjs
 +
 
 +
Source code in /storage/big1/lml/jac/was_invitro/Projects/cspat-volume
 +
 
 +
</nowiki>
 +
== Image Analysis ==
 +
===  Corecell.pl  ===
 +
<nowiki>
 +
 
 +
Takes a 3d image and an outline and keeps all data pts within the specified distance
 +
of the outline (removes the "core" of the cell). 
 +
Also produces a file named in_image_H.histo. See help for histogram2 about that.
 +
If in_image2.i2i out_image2.i2i are specied, in_image2.i2i is also cored
 +
(and histogram2 is applied to the pair in_image.i2i and in_image2.i2i when producing in_image_H.histo).
 +
 
 +
Written to let Ronghua only keep BK and Ryanodine label near the plasma membrane.
 +
Note: touching voxels may have a distance of .5 voxels, and voxels at the same
 +
      position may have a distance of -.5 (??)
 +
Note: pixels may be kept near the end of the cell, since they have close distance to the end
 +
      (rather than to the plasma membrane. Might -Z # # option to histogram2 fix this?).
 +
Note: there might be problems if boundary.pts touches the edge of the image.
 +
 
 +
 
 +
Corecell.pl [options] in_image.i2i boundary.pts out_image.i2i [in_image2.i2i out_image2.i2i]
 +
 
 +
options:
 +
-interp interpolate missing z slices in boundary.pts before using it.
 +
zslices must be in ascending order in boundary.pts.
 +
contours must all be drawn in the same direction (i.e., all clockwise or all counter-clockwise).
 +
The name of each object is changed to "nuke".  A maximum of 1000 zslices is allowed.
 +
-shrink=#  shrink boundary.pts by this number of pixels before using it
 +
-expand=#  expand boundary.pts by this number of pixels before using it
 +
-zrange=#:#    only examine z slices (1-indexed in this range)
 +
-dist=#: keep all voxels within this distance of the boundary, default = 2
 +
-dx=#          pixel size x size (default = 1. all that really matters is relative x,y, and z size)
 +
-dy=#          pixel size y size (although if you change this then that may affect -dist)
 +
-dz=#          pixel size z size
 +
-dontcapends    first and last outline slices aren't capped (removes -c from the call to histogram2).
 +
        typically you'd use this if the outlines are circular cross-sections of a cylindrical
 +
cell (ie, z is along the length of the cell).  If the outlines are along the length
 +
of the cell (ie, cell is in the xy plane, as Jeff likes to do), then you wouldn't use this option.
 +
-verbose print out extra info to stderr as it goes along
 +
-vverbose very verbose
 +
-keepfiles keep all intermediate files (all begin with _)
 +
 
 +
example:
 +
  #cell's length is in the xy plane, need to flip it to get cross sections to outline with planimeter3
 +
  transp -YZ im.i2i im_YZ.i2i
 +
planimeter3 -I im_YZ.i2i -R im_YZ.pts
 +
  transp -YZ im2.i2i im2_YZ.i2i
 +
  #cell's original zspacing = 250 nm, but x and y are 80, so relative spacing is about 3:1, after transp
 +
  #it is the y direction that has the 250 nm spacing.
 +
Corecell.pl -dy=3 -dist=3 -interp im_YZ.i2i im_YZ.pts im_YZ_near_pm.i2i im2_YZ.i2i im2_YZ_near_pm.i2i
 +
dave -T 1 YZ -T 2 YZ -z 3 -I im_YZ_near_pm.i2i -I im2_YZ_near_pm.i2i
  
 +
see also: shrink_wrap (to apply prior to Corecell.pl to get a better starting surface)
 
   
 
   
−
  see also:
+
</nowiki>
−
   channel_current2
+
=== count_2d_objs2.pl  ===
−
   /home/lml/krypton/Zhuge/BKandRyr/analysis6/run12
+
<nowiki>
−
   /home/lml/krypton/Huang/Kth_pt.m
+
run countobjs on a 2D time series
−
   p. 637 of Cressie.  
+
creates temporary files _temp.i2i and _temp.stats in the current directory.
 +
note: scale and black level forces each time point to be rescaled the same and
 +
      therefore to be thresholded the same.
 +
file.stats will have data ordered as:
 +
time number_of_objects mean_iod sd_iod mean_size sd_size
 +
 
 +
count_2d_objs2.pl [options] image.i2i file.stats
 +
 
 +
options:
 +
   -tstart=# first time point to examine, default = 1
 +
  -tstop=# last time point to examine, default = last z slice.
 +
  -min=# minimum size object to allow, default = 1
 +
  -max=# maximum size object to allow, default = 100000
 +
  -thresh=# threshold, default = 0
 +
  -scale=# scale, default = 1
 +
  -black=# black, default = 0
 +
  -allstats=filename print info about each object to _temp.stats (otherwise just summary stats from each t printed)
 +
and rename it "filename"
 +
   -outimage=image.i2i see -o option in countobjs.
 +
   -Outimage=image.i2i see -O  option in countobjs
 +
   -Qimage=image.i2i see -Q option in countobjs, note: id numbers will restart from 1 on each slice.
 +
 
 +
  -verbose
 +
  -dummy
 +
  -noclean
  
 
   
 
   
−
===  coloc.pl  ===
+
</nowiki>
 +
===  distance2surface_overtime.pl  ===
 +
<nowiki>
  
−
calls coloc3way with a bunch of different thresholds. Creates an output file which can be
+
Finds the distance (in pixels) of a specified pt (over time) from the nearest pixel
−
graphed with VennDiagram.m.  
+
greater than 0 in 3Dimage.i2i (or greater than -threshold).  Alternatively, finds
 +
the distance to the boundary surface specified with the -surface option.
 +
The coords is as produced by track_pts (x y z otherstuff).
 +
Written to let Andrea take a tracked gene in the nucleus and find its
 +
distance to the nuclear envelope over time.
 +
Note: touching voxels may have a distance of .5 voxels, and voxels at the same
 +
      position may have a distance of -.5 (??)
  
−
coloc.pl im1.i2i im2.i2i im3.i2i  > out.coloc3way
+
distance2surface.pl [options] 3Dimage.i2i specifiedpt.coords outfile.dist
  
 
options:
 
options:
−
-time_vs_thresh just generate a file which shows which times correspond to which thresholds. images can be dummy names.
+
-surface=boundary.pts find distance to the surface defined by boundary.pts
 +
  rather than the distance to nearest >0 voxel 3Dimage.i2i
 +
-shrink=#  shrink boundary.pts by this number of pixels before using it
 +
-expand=#  expand boundary.pts by this number of pixels before using it
 +
-zrange=#:#    only examine z slices (1-indexed in this range)
 +
-threshold=#    threshold 3Dimage.i2i (voxels <= threshold set to 0, > threshold set to boundary).
 +
-pixel=# pixel size (in x and y) in nm
 +
-verbose print out extra into to stderr as it goes along
 +
-vverbose very verbose
 +
-keepfiles keep all intermediate files (all begin with _)
  
−
-t1=#:#:# threshold one should go from # to # by #. default = 800:1200:50
+
−
-t2=#:#:# threshold two should go from # to # by #. default = 800:1200:50
+
</nowiki>
−
-t3=#:#:# threshold three should go from # to # by #. default = 2000:4000:200
+
=== extract_area2.pl  ===
−
-numthresh=#1:#2:#3  ignore the third number in the -t1, -t2, and -t3 options (or in the default) and instead calculate a
+
<nowiki>
−
  threshold step size so that #1 thresholds are applied to im1.i2i (dat image)
+
Can't use 'defined(@array)' (Maybe you should just omit the defined()?) at /home/lml/krypton/bin/extract_area2.pl line 85.
−
  #2 thresholds are applied to im2.i2i (ctx image), #3 thresholds are applied to im3.i2i (rin image).
 
−
  e.g., -numthresh=5:10:10 will apply 5 different thresholds to im1.i2i, and 10 thresholds to im2.i2i and im3.i2i.
 
−
  Using this option lets me insure that each call to coloc.pl will produce exactly the same number of data points;
 
−
  this may make the statistical comparison between cells easier.
 
  
−
-Melikian=file in addition to creating output for VennDiagram, create text file with the coloc percentages
+
</nowiki>
−
Dr. Melikian is interested in. Graph via xmgr -nxy. See header in file for coloc info.
+
===  findpaths_exo4.pl ===
−
For this to work the three image files MUST be specified in the order 405 594  488 (i.e., rin.i2i ctx.i2i dat.i2i)
+
<nowiki>
−
-Melikian2=file Same as -Melikian, but dat (488) is image 1, ctx (594) image 2, and rin (405) image 3
+
 
−
-area=file.xyr also print out Melikian data as xyr where r is proportional to the number of pixels involved
+
this program takes output from exo4.
−
(i.e., the denominator used when calculating the percent colocalized).
+
similar to findpaths_exo3.pl, but parses exo4 output rather than exo3 output.
−
prints to "file.xyr".  Must also specify either -Melikian or -Melikian2
+
 
−
xmgrace -graphtype fixed -settype xyr file.xyr
+
usage:
−
-matlab=file:#  like -area but as columns so easy to read into matlab: t p1 n1 p2 n2 p3 n3 p4 n4
+
   findpaths_exo4.pl file1.out
−
# is the max n value (ie, all n values are scaled so the max over n1,n2,n3,n4 is #).
+
 
−
Since p1,p2,p3,p4 are percent, # is typically set to a small part of that, e.g. .05.
+
options:
−
If # is negative then abs(#) is just used as a scale factor (so -1 would just keep the
 
−
n values as the number of voxels). Graph in matlab with line width of 2*n1:
 
−
matlab   # import data=(tdim,9)
 
−
>> lowy = data(:,2) - data(:,3); hiy = data(:,2) + data(:,3);  x = data(:,1);
 
−
>> area(x,hiy,'FaceColor',[1 0 0],'EdgeColor','r','LineWidth',1); hold on;
 
−
>> area(x,lowy,'FaceColor',[1 1 1],'EdgeColor','r','LineWidth',1);
 
−
Must also specify -Melikian or -Melikian2.
 
  
−
-debug
+
which paths:
−
-verbose
+
  -prestring="# debugpath 3 (102,156,-1):"  pull out paths that begin with this. default is the null string.
 +
    : null string paths should also be saved_paths later.
 +
    : "# debugpath .*:"  will pull out all the debugpaths (. matches any char and * means
 +
    : number of them)
 +
  -saved_paths     :  all the saved paths (within -zrange if that is also specifed)
 +
  -debugpaths     :  all the debug paths.  These options are mutually exclusive.
 +
  -zrange=minz:maxz     :  only print tracks which start within the z range specified (1-indexed, inclusive)  
  
−
example:
+
what to do with the paths:
 +
  -saved_labels=filename    :  implies -saved_paths. saves labels for each path into the specified filename. This
 +
    :  file can be read into play so that labels are visible at a location.
 +
for rpts file:
 +
  -rptsfile=track.rpts      :  create an rpts file (initially empty). use -rptstracks and -printfusion to put stuff in it.
 +
  -rptstracks     :  put paths (tracks) into the rpts file specified with -rptsfile    
 +
  -printfusion=#     :  print box (of width #) around fusion events to rptsfile. must us -rptsfile option with this.
 +
    :  note: -debugpaths do not have valid fusion times.
 +
  -pathcolor=#     :  instead of -1
 +
  -boxcolor=#     :  color of fusion box. instead of -2
  
−
coloc.pl im1.i2i im2.i2i im3.i2i > coloc.data
+
for xmgr file:
−
coloc.pl -time_vs_thresh im1.i2i im2.i2i im3.i2i > coloc.times
+
  -trackxmgr=track.xmgr    :  create an xmgr file of the tracks produced by the program "exo".
−
matlab
+
  -truetime     :  rather than use z value as "time" calculate time based on the "burst protocol"
−
>> VennDiagram('coloc.data','percent')  # and manually look at coloc.times while doing this to convert times to thresholds.
+
    :  of 60 seconds acquisition at 1Hz followed by 3 seconds at 20HZ, repeatedly.  
 +
  -peak     :  rather than print avg (iod/size), print the peak pixel value (ie, value of maxima).
 +
  -finalfit     :  rather than print avg, print the best fit curve (based on final fit parameters).
 +
  -initfit     :  rather than print avg, print the init fit curve (based on initial fit parameters - for debugging).
 +
  -distance     :  rather than print avg, print the distance from the first pt on the curve.
 +
  -maxtracks=#     :  for -trackxmgr, only print out this many tracks.
 +
  -tracks=start:skip     :  for -trackxmgr, only print tracks starting at track "start" (1-indexed) and
 +
    :  skip "skip" paths beforing printing another one (e.g., print every skip paths).
 +
    :  this lets you print a subset of paths which are still spread over the entire z range
 +
    : to make visualization easier. skip = 0 means don't skip any.
 +
misc:
 +
  -verbose
 +
  -stationary_time=filename : print time each path is stationary prior to fusion to specified file
  
−
see also:
+
example:
−
~/krypton/packages/fiji/Fiji.app/plugins/jacob_.jar  - more traditional colocalization measures
+
  exo4 -d 120 -o adp12273_ids.i2i -V -w 10 -b 2 adp12273_c1_z1201_1320_masked.i2i > & adp12273_c1_z1201_1320_masked.out
−
coloc_tseries2
+
  play -min 0 adp12273_ids.i2i    # maxima of each region should be blue
−
colocalizeL
+
  findpaths_exo4.pl -trackxmgr=adp12273.xmgr -truetime adp12273_c1_z1201_1320_masked.out
 +
  xmgr adp12273.xmgr    #clicking on each set brings up its legend info which has its
 +
        #starting coord (1-indexed I think) and value.
 +
  findpaths_exo4.pl -rptsfile=adp12273.rpts -rptstracks -printfusion=4 adp12273_c1_z1201_1320_masked.out
 +
  addlines adp12273_c1_z1201_1320_masked.i2i adp12273.rpts - | play -P -1 R -P -3 G -
  
 +
see also:  ~lml/krypton/Huang/TIRF4/graph_exo3.csh
 
   
 
   
−
===  probcircle.pl  ===
+
</nowiki>
−
  given a sphere of diameter D, what is the prob of oberving a disk of radius r when a random plane intersects it?
+
===  get_find_vesselness_results.pl  ===
−
  prints to stdout: r prob(r) 1/prob(r)
+
<nowiki>
 +
  Can't use 'defined(@array)' (Maybe you should just omit the defined()?) at /home/lml/krypton/bin/get_find_vesselness_results.pl line 115.
  
−
   usage:  
+
</nowiki>
−
      probcircle.pl [options]  
+
===  mask_and_coloc.pl  ===
 +
<nowiki>
 +
Takes a threshold, x and y translation for vinculin image, a pts file, and two images, and creates a mask
 +
and then calculates colocalization. See README.
 +
 
 +
mask_and_coloc.pl -x=# -y=# -thresh=# -pts=file.rpts -vin=vin.i2i -trip=trip.i2i
 +
 
 +
options:
 +
  -tx=# translate by a max of -tx when calculating other colocalization numbers, default = 25
 +
  -ty=#
 +
  -step=# step size for other translations, default = 5
 +
  -noclean don't clean up (ie, don't remove) intermediate files created during this analysis
 +
  -overlap=file   just calculate overlap3, reusing the other files, put result into file.
 +
 
 +
 +
</nowiki>
 +
===  mask_gedmat_images4.pl  ===
 +
<nowiki>
 +
  mask_gedmat_images4.pl  -check_intersect  added.
 +
determines when outlines intersect and sets the aerr (area error flag) in the
 +
output file. eventually it will calculate areas a different way when that happens
 +
rather than combining masked images (see ~/krypton/Corvera/Raz/README for 10/18/13). not done yet ...
 +
 
 +
  Examines all the zip files (produced within fiji when outlining regions) in the current directory. These have
 +
  the boundary of the explant, day7 growth, and day11 growth.
 +
  note: this program assumes that all the data in the directory is from the same patient and the same tissue type.
 +
 
 +
  It recalculates the area measures by only including that part of the day7 and day11 growth which is between the explant
 +
  center and going towards the center of the well; area from the explant center going away from the center of the
 +
  well (ie, growing towards the well wall) is excluded. By doing this, we hope to minimize the
 +
  effect of growth (as measured by area) being stopped by the well wall when the explant is near the well wall. 
 +
  Output is written to stdout. The comments there explain the fields. area_mex is the old value, area_mex_border is the new.
 +
 
 +
  This is done by finding all the *_Area.zip files in the current directory, pulling out the outlines in those files
 +
  and using that information to recalculate areas.  It also needs to find the associated "combined" image file
 +
  (*_combined.i2i) to get image size and hence the position of the center of the well) and the fiji results file
 +
  (*_results.txt, so the numbers produced here can be easily compared with them).
 +
 
 +
  note: The algorithm assumes that the area NEVER DECREASES with time, and that the explant region is always surrounded
 +
by the day7 region which is surrounded by the day11 region. The latter assumption is sometimes invalid if
 +
the day 11 images were shifted relative to day 7 images; but that is usually ok.
 +
(if necessary, you can manually change the "bad_data" field in the output file to 1 and then use
 +
-cond in gedmat_to_xy.pl to eliminate those stats from further analysis.)
 +
It also assumes the smallest area in each zip file is the original explant area, the next largest area is
 +
called day7 area and the largest area (if 3 are present) is assumed to be day 11 area.
 +
Sometimes when growth is about 0, the outlines on, eg, day7, may have slightly less area than the original
 +
explant, so the outlines might be mis-ordered then. [actually it may read the correct day off the name of
 +
the i2i image now].
 +
 
 +
  note: If only 2 areas (ie, two .roi files) are present in the zip file, they are assumed to be for
 +
the explant and day7 (not day11).
 +
 
 +
  note: This program assumes the center of the combined image (usually 4 well images are combined into one) is the
 +
center of the well. see combine_images.pl. It also assumes the combined image name ends in combined.i2i
 +
(see -combined).
 +
 
 +
  note: some of the scripts I use to then analyze these results will multiply the area by 2, to get a "true" estimate
 +
  of growth; since the value produced by mask_gedmat_images3.pl will be (roughly) half of the real growth rates.
 +
 
 +
   usage:
 +
mask_gedmat_images3.pl [options]  
  
 
   options:
 
   options:
−
-D=# default = 1
+
-tmpdir=fullpathname directory to use for temporary files which are created, eg: -tmpdir=/home/lml/krypton
−
-just_prob don't print the third column
+
 
 +
-zip_file=file.zip
 +
-zip_file=file.zip:file2.zip:...:filen.zip
 +
only examine the one specified zip file, not all those in the directory
 +
(If the second form is used, you can specify as many as you want.)
 +
note: you may still need to specify -zip_ending so that the ending of this file can be stripped
 +
      off correctly (so the -combined_ending and -results_ending can then be added correctly).
 +
-zip_ending=stuff e.g., -zipending=_area.zip  .  All files which end in this will be analyzed; default = _Area.zip
 +
-skip=file.zip[:file2.zip...]  list of zip files (without the full path) to skip.
 +
 
 +
-results_ending=stuff e.g., -results_ending=_results.txt  .  This is what the -zip_ending should be replaced by
 +
to get the name of the associated fiji results file.  note the _ should be included here
 +
if it was included in -zip_ending.
  
−
  examples:
+
-combined_ending=stuff  e.g., -combined_ending=_combined_reduced_.i2i  . default = combined.i2i.
−
      probcircle.pl | xmgrace -free -pipe -nxy -
+
This is any string which helps identify the last part of the combined image filename.
 +
-combined_scaling=#:# e.g., -combined_scaling=2:2 if reduceima -bin 2 2 was applied to get _combined_reduced_.i2i.
  
−
  see also: p. 249 stereology_book.pdf
+
-scale=#:# distance and area (eg, -scale=2:4 if reduceima -bin 2 2 was used) scaling. This will multiply
−
    /home/lml/krypton/packages/fiji/stereology_lml.template
+
the area stored in the fiji file and the area calculated via my mask by the second number.
−
    /home/lml/krypton/Corvera/Raz/stereology/plot_results.pl
+
The first number will multiply the distance of the explant from the center of the image.
−
    /home/lml/krypton/facil/cross_sections_of_sphere
+
This option is just here to handle 001_BAP and 002_PRO datasets. They were analyzed in fiji
 +
with the images reduced by a factor of 2 in x and y. So the fiji areas produced will be low
 +
by 4x, as will my areas.  This option compensates for that.  See ~/Corvera/Raz/README for 10/11/13.  
  
−
   
+
-check_intersect check if outlines intersect with the explant's outline. If so, set an error flag on stdout printout
−
===  compare2dist  ===
+
and print info to stderr. eventually will just use area numbers rather than masking area in this situation.
 +
that is not done yet ...
  
−
This program calculates whether two amplitude distributions are the same.
 
−
It was written to examine Rhonghua's STOCS data to determine whether the increase
 
−
in observed STOC frequency could be explained by an increase in STOC amplitude
 
−
(since then more events above threshold would be observed). 
 
−
datafile2 should be the one with larger amplitudes (so we hypothesize it has fewer missing
 
−
events if it is just a version of datafile1 shifted higher).  It will get shifted down
 
−
by a range of shifts (-s option) and compared with datafile1.
 
−
This routine also compares the temporal interarrival time distribution of each datafile with
 
−
that predicted by a poisson process and with the other datafile; the temporal calculations
 
−
ignore all missing data issues and shifting issues for now.
 
  
−
This program reads data in the form:
+
-save_day[=directory]
−
    time calcium prior_baseline post_baseline amplitude
+
saves the final binary mask for each days growth (eg, day7 growth minus the explant and minus the part growing towards the wall)
−
      where the values are
+
into a 3d mask image (patientid_tissue_day#_mask.i2i).  This image will be reduced down in xdim and ydim from the
−
      time:           time of STOC
+
original.  patientid_tissue_day#_.play_labels will also be created (labels which well each z slice comes from).
−
      calcium:        SR calcium level
+
This is mostly for quality control purposes.
−
      prior_baseline: amplitude just prior to STOC
+
In addition, patientid_tissue_day#.rpts will be created (similarly reduced down in size) (coords may be off by 1?).
−
      post_baseline:   amplitude just after STOC
+
View the file via: addlines maskim.i2i maskim.rpts -  |play.FC14 -P -1 R -labels maskim.play_labels -
−
      amplitude:       amplitude amplitude of STOC (not relative to baseline)
+
  or via: play_results3.pl
−
Data is expected to already be sorted by time (smallest to largest).
+
patientid_tissue_day#_orig.i2i will be the original images (also reduced in xdim and ydim, and only one 2d slice
−
comment lines must begin with a #
+
from each original 3d image).
 +
note: older versions of play may need the comment line at the top of maskim.play_labels removed.
 +
note: if you want images to be in another directory, specify a full pathname, eg -save_day=/home/lml/krypton (without last /).
 +
note: any existing file with these names will be OVER-WRITTEN.
 +
note: this option creates lots of temporary files (in tmpdir) of the form __*.i2i, any existing
 +
      files that match this will be DELETED.  
 +
see also: montage
  
−
Usage: compare2dist [options] datafile1 datafile2
+
-just_check just check whether each zip roi outline has the correct number of associated image files.  
−
options:
+
eg, if the zip file has 3 outlines (explant, firstday, secondday) there should be two i2i files with DAY# in their name.
−
    -t #: only include data with an amplitude > #. The default
+
prints to stderr info about those which don't have the correct number. does nothing else.
−
is to assume a 0 threshold. If you have acquired the data
+
-verbose some extra info to stdout
−
using some thresholding criterium (eg, only peaks above
+
-debug prints some tracing info to stderr
−
10picoamps are counted) then you should provide that number
+
-print_fname print name of each zip file as it is being analyzed, to stderr.
−
here, even if all your data is above this (ie, you've applied
+
-dlevel=# extra debugging. the bigger the number the more the printout. # is an integer starting at 1.
−
the threshold in your identification phase). This is because
+
-dlevel=1 will print a trace of commands to execute to mimic (parts of) this perl script.
−
we need to know it when we shift amplitudes of the second
+
-noclean don't remove the temporary files which were created
−
distribution to see if it could match the first distribution.
+
the files are: _ones.i2i _explant.rpts _day11.rpts _day7.rpts _excluded_region.rpts
−
    -b: subtract average baseline from amplitude prior to use (ie, prior
+
      _explant.i2i  _day11.i2i _day7.i2i _day7_noex.i2i _day11_noex.i2i _day7_noex_halfex.i2i _day11_noex_halfex.i2i
−
to any thresholding or intensity shifting of data).
+
  (if multiple zip files were analyzed, only the files from the last one will be left, since they get reused)
−
    -s delta max: shift datafile2 amplitudes down by a maximum of max, in
+
-dont_recalc only for diagnostics. does NOT restrict the analysis to just the growth away from the wall.
−
increments of delta, to see if it will then match datafile1.
+
with this specified this entire program should just reproduce the numbers we already
−
  -p #: our guess of the percentage of data missing from datafile2 because it was
+
have in *Results.txt since that uses the entire area. ie, this make the program a no-op.
−
below our threshold/detection criteria (a # from 0 to 1).
 
−
This plus the amount of datafile2 shifted below threshold determines
 
−
our estimate of the amount of data missing from the shifted datafile2 and
 
−
the unshifted datafile1.  The larger this number is, the more likely
 
−
the two curves will appear similar (since differences in actual data
 
−
examined will be proportionally smaller differences, and we are assuming
 
−
all missing data matches exactly, but see -m below). Default = 0.000000
 
−
    -m: assume all missing data maximally MISmatches, this would probably be
 
−
a worst case scenario, so that if the KS statistic says the distributions
 
−
could be the same even with the -m option, its probably true. But I'm
 
−
not certain of this, since it really depends how this affects the  
 
−
relative spacing between the cumulative distributions. I'd have to
 
−
think about it some more.
 
  
−
see also:test_distr, poisson
+
  see also:
−
This code is in ~lml/vision/Rhonghua/STOCS
+
/home/lml/krypton/Corvera/Raz/README
−
#/home/lml/krypton/bin/compare2dist
+
analyze_gedmat3.pl a script which calls mask_gedmat_images3.pl for all of Raz's data. Also versions 4,5,6, and 7.
 +
play_results3.pl
 +
raz_rates.pl
 +
mask_gedmat_images.pl
 +
mask_gedmat_images2.pl
 +
/storage/big1/raz/GEDMAT/
 +
ReadROI.pl
 +
extract_area.pl
 +
combine_images.pl
 +
gedmat_to_xy.pl
 +
/home/lml/krypton/Corvera/Raz/analyze_gedmat.pl
 +
chres_rpts.pl
 +
  filter_rptsinz.pl
  
−
=== compare2gaussian  ===
+
  examples:
−
incorrect command line, need 1 data files.
+
cd /storage/big1/raz/GEDMAT/006_JTH/OM
−
This program calculates whether a given set of data could be samples from a known mixture of Gaussians.
+
 
−
datafile should have one data point per line. comment lines begin with #.
+
# generate 006_JTH_OM.areas with all the data (area_mex_border is the number which should replace farea)
−
Output statistics to stdout.
+
# also produce 006_JTH_OM_7masks.i2i and 006_JTH_OM_11masks.i2i so I can check on the validity of the algorithm.
 +
# place diagnostic mask images in /home/lml/krypton/Corvera/Raz/output and the output data file in
 +
# /home/lml/krypton/Corvera/Raz (note: all directories must already exist).
 +
mask_gedmat_images.pl -tmpdir=/home/lml/krypton/Corvera/Raz/output  -zip_ending=_Area.zip -results_ending=_Results.txt
 +
-combined_ending=combined.i2i -days=7:11
 +
-save_day=/home/lml/krypton/Corvera/Raz/output
 +
>& /home/lml/krypton/Corvera/Raz/006_JTH_OM.areas
 +
 
 +
# look at the masks produced:
 +
cd /home/lml/krypton/Corvera/Raz/output
 +
      play_results3.pl 006_JTH_OM_day7
 +
or:
 +
fgrep -v # 006_JTH_OM_11masks.play_labels > tmp.play_labels  # eliminate the comment line, play doesn't like those.
 +
play -labels tmp.play_labels 006_JTH_OM_7masks.i2i
 +
 
 +
# compare the full areas produced which what fiji did (as a check). only pull out the good data. they should be very similar:
 +
cd /home/lml/krypton/Corvera/Raz
 +
gedmat_to_xy.pl -cond='if (($date eq "date_sure")&&($perr==0)&&($aerr==0)) {1;} else {0;}'
 +
-xcoord='$area' -ycoord='$farea' 006_JTH_OM.areas | xmgrace -free -pipe
 +
 
 +
# now grab the data we really want. compare the old way, with the area when the growth half near the well border is masked off:
 +
gedmat_to_xy.pl -cond='if (($date eq "date_sure")&&($perr==0)&&($aerr==0)) {1;} else {0;}' 
 +
-xcoord='$area_mex' -ycoord='$area_mex_border' 006_JTH_OM.areas | xmgrace -free -pipe
 +
 
 +
# it may be hard to see some of these images as they are very large, do it this way (note the -L to keep the -1 boundary values):
 +
addlines _ones.i2i _excluded_region.rpts - | reduceima -bin 3 3 -L - -|play -P -1 R -P -2 G -P -3 B -P -4 C -P -5 Y -
  
−
Usage: compare2gaussian [options] datafile
 
−
options:
 
−
      -t #: only include data with an amplitude > #. The default is to include all data.
 
−
  -g # amp1 mean1 sd1 ... :
 
−
The Gaussian mixture model the data should be compared against (using a KS statistic).
 
−
# is the number of Gaussians, this is followed by the amplitude, mean, and sd of
 
−
each Gaussian. The default is one Gaussian with mean = 0 and sd = 1.
 
−
You can get these values using the fitgaussians2 program.
 
−
  -v: verbose. Also prints the observed (first) and theoretical (second) curves to stdout.
 
−
  -T: test. runs through x = 0 to 10 and prints out cumulative prob. exits.
 
  
−
See Also: fitgaussians2, fit2gaussians, fit3gaussians
+
# run with some diagnostics:
 +
cd /storage/big1/raz/GEDMAT/006_JTH/OM
 +
/home/lml/krypton/bin/mask_gedmat_images.pl -tmpdir=/home/lml/krypton/Corvera/Raz -zip_ending=_Area.zip
 +
        -results_ending=_Results.txt -combined_ending=combined.i2i -days=7:11
 +
        -save_day=/home/lml/krypton/Corvera/Raz/006_JTH_OM_7masks.i2i    > /home/lml/krypton/Corvera/Raz/006_JTH_OM.areas
 +
# play.FC14 allows labels to have comment lines (first character of line is a #)
 +
cd home/lml/krypton/Corvera/Raz
 +
play.FC14 -labels 006_JTH_OM_11masks.play_labels 006_JTH_OM_11masks.i2i
 +
chres_rpts.pl -sx=.33 -sy=.33 -truncate -min=1:1 output/_day11.rpts > _day11_bin3.rpts
 +
chres_rpts.pl -sx=.33 -sy=.33 -truncate -min=1:1 output/_day7.rpts > _day7_bin3.rpts
 +
chres_rpts.pl -sx=.33 -sy=.33 -truncate -min=1:1 output/_explant.rpts > _explant_bin3.rpts
 +
reduceima -bin 3 3 output/_day11_noex_halfex.i2i - | addlines - _day11_bin3.rpts - | addlines - _day7_bin3.rpts - |
 +
addlines - _explant_bin3.rpts - | play -P -1 R -
 +
addlines /storage/big1/raz/GEDMAT/006_JTH/OM/D07_20130524_GEDMAT_006_JTH_OM_DAY7_combined.i2i output/_day11.rpts - |
 +
addlines - output/_day7.rpts - | addlines - output/_explant.rpts - | reduceima -bin 3 3 -L - - | play -P -1 R -
  
−
Source code is in ~lml/krypton/Lefkowitz
 
−
#/home/lml/krypton/bin/compare2gaussian
 
  
−
===  compare_dist2  ===
+
# do a bunch of stuff
 +
analyze_gedmat3.pl # apply this perl script to lots of data
 +
cd output
 +
      play_results3.pl 006_JTH_OM_day7  # look at some of it
 +
raz_rates.pl # generate stats
 +
  
−
This program calculates whether two distributions are the same using chi-square test (unless -k).
 
−
It was written to examine the distribution of sizes of Ryanodine Receptor objects
 
−
to determine if they are randomly distributed. datafile1 should be the reference
 
−
data (gotten from many simulations or from a theoretical formula), datafile2
 
−
should be the real data (max number of data values = 100000).
 
  
−
This program reads data which is just a list of sizes of objects seen (one per line).
+
  source code: in /home/lml/krypton/bin/mask_gedmat_images.pl
−
Comment lines must begin with a #
 
−
Floating pt numbers are truncated to integers for bin number (max bin number = 50000).
 
  
−
Usage: compare_dist2 [options] datafile1 datafile2
+
−
options:
+
</nowiki>
−
  -k perform K-S test instead (I'm not certain the numbins I use to calculate this is correct).
+
===  mask_gedmat_images4_new.pl  ===
−
  -a also print the histograms produced
+
<nowiki>
−
  -b also print the normalized histograms (total count datafile2 scaled to the same count as in datafile1,
+
  mask_gedmat_images4_new.pl
−
  bins with 0 removed), this is only done if -k not specified (ie, only if performing chi-square test).
+
The easiest way to call this is via analyze_gedmat9.pl, see its help.
−
  -c also print the cumulative prob distributions (only done if -k also specified)
+
   
−
  -r # #: only analyze sizes within the specified range (inclusive)
+
This version has the following changes compared with mask_gedmat_images3.pl:
−
  -m #: cap the maximum number of counts for any size in datafile1 to #
+
 
−
  -M #: cap the maximum number of counts for any size in datafile2 to #
+
1. @stats = &outline_stats(@outline) was used to calculate explant center (for masking) but
−
  -z: for chi-square keep zero entry bins (chi square only correct if this option is NOT used)
+
        the meanx and meany are just the mean of the outline, it is NOT a center of mass. usually ok for a compact explant.
−
so why is this option here?  if you just really want to use this program to do
+
  sub calc_com() is now used instead to calculate the true center of mass.  
−
histogram normalization (-b), and don't care about the stat. xmgrace running avg
+
-use_explant_outline will cause the OLD method to be used instead.
−
will produce strange results if you don't use -z, ie, if data not equally spaced.
+
 
−
  -v verbose
+
2.  bounding box was used to calculate area for sorting the outlines temporally (from smaller to larger area).  
−
  -V very verbose
+
      but when outlines intersect bounding box can at times NOT be a good surrogate for area.
−
example:
+
  $roi_area[$r] = ($maxx-$minx)*($maxy-$miny); 
−
      create a simulated distribution and compare it with real restored data (real_ryrr.i2i, run5 does this 100 times):
+
  handled these two issues by writing sub calc_com().
−
  cluster -wsize 10 43 25 -cylinder 0.001 3 -psize 0.08 0.08 0.25 -mindist 0.030 -numryr 500 -image ryr.i2i
+
-use_bbox_area will cause the OLD method to be used instead.
−
  blur3d -S 40000 -P ryr.i2i psf.i2i ryr_b.i2i
+
 
−
  noise -c RCA ryr_b.i2i ryr_bn.i2i
+
 
−
  setenv myloc /home/lml/krypton/Zhuge/Ryr_clusters
+
note: fiji areas are already sorted smallest to largest when I first bring them in (extract_area2.pl does that):
−
  setenv eprloc /storage/big1/epr/queues
+
  $prev_results = `extract_area2.pl -justfiles=$resultsfile`
−
  echo "  /storage/big1/epr/run_epr -V -log -NSC 4 -sm 1e-05 -it 5000 -sc 0.025 -co 0.001
+
  so I should do not need to use the sorted roi indices on them (so I've been doing this correctly). but the rpts are
−
-psf ${myloc}/psf.i2i ${myloc}/ryr_bn.i2i ${myloc}/ryr_bnr.i2i " > /storage/big1/epr/queues/standard/lml.0
+
  being sorted based on bounding box, and fiji areas based on the true area in the Results.txt file. So the ordering
−
  while (1==1)  if ( ( ! -e ${eprloc}/standard/lml.0 ) && ( ! -e ${eprloc}/running/lml.0 ) ) then break endif sleep 60s end
+
  may be different at times. No change was made to this.
−
  countobjs -k -s 5 300 -t 4500 4500 sim_ryr.objs ryr_bnr.i2i
 
−
  cull -s -c -2 < sim_ryr.objs > sim_ryr.stats
 
−
  countobjs -k -s 5 300 -t 4500 4500 real_ryr.objs real_ryrr.i2i
 
−
  cull -s -c -2 < real_ryr.objs > real_ryr.stats
 
−
  compare_dist2 -b sim_ryr.stats real_ryr.stats  >& chi_ryr
 
−
  compare_dist2 -k -b sim_ryr.stats real_ryr.stats  >& ks_ryr
 
−
  tail chi_ryr ks_ryr
 
−
  xmgrace chi_ryr; xmgrace ks_ryr
 
  
−
see also: cluster, run5
+
  mask_gedmat_images4.pl  -check_intersect  added.
−
This code is in /home/lml/krypton/Zhuge/Ryr_clusters/
+
determines when outlines intersect and sets the aerr (area error flag) in the
−
#/home/lml/krypton/bin/compare_dist2
+
output file. eventually it will calculate areas a different way when that happens
 +
rather than combining masked images (see ~/krypton/Corvera/Raz/README for 10/18/13). right now only sets the error flag.
  
−
===  correlationL ===
+
  Examines all the zip files (produced within fiji when outlining regions) in the current directory. These have
−
/home/lml/krypton/bin/correlationL
+
  the boundary of the explant, day7 growth, and day11 growth.
 +
  note: this program assumes that all the data in the directory is from the same patient and the same tissue type.
 +
 
 +
  It recalculates the area measures by only including that part of the day7 and day11 growth which is between the explant
 +
  center and going towards the center of the well; area from the explant center going away from the center of the
 +
  well (ie, growing towards the well wall) is excluded. By doing this, we hope to minimize the
 +
  effect of growth (as measured by area) being stopped by the well wall when the explant is near the well wall.  
 +
  Output is written to stdout. The comments there explain the fields. area_mex is the old value, area_mex_border is the new.
  
−
This program plots intensities from image1 vs intensities from
+
  This is done by finding all the *_Area.zip files in the current directory, pulling out the outlines in those files
−
image2. image1 on the x axis, image 2 on the y axis. Intensities
+
  and using that information to recalculate areas. It also needs to find the associated "combined" image file
−
in both images are rescaled 0 to the size of the output image
+
  (*_combined.i2i) to get image size and hence the position of the center of the well) and the fiji results file
−
(default = 512).  The intensity at each pixel in the output
+
  (*_results.txt, so the numbers produced here can be easily compared with them).
−
image is the number of (image1,image2) voxels with that particular
+
 
−
intensity combination.
+
  note: The algorithm assumes that the area NEVER DECREASES with time, and that the explant region is always surrounded
−
The intent is to be able to look at the output image and get a sense
+
by the day7 region which is surrounded by the day11 region. The latter assumption is sometimes invalid if
−
for whether there is a correlation between intensities
+
the day 11 images were shifted relative to day 7 images; but that is usually ok.
−
in the two images.  If so, perhaps a least squares line
+
(if necessary, you can manually change the "bad_data" field in the output file to 1 and then use
−
fit should be done to see how well one images correlates with the
+
-cond in gedmat_to_xy.pl to eliminate those stats from further analysis.)
−
other, or -s option can be specified.
+
It also assumes the smallest area in each zip file is the original explant area, the next largest area is
−
The minimum, maximum, and scale factors are printed to stdout.
+
called day7 area and the largest area (if 3 are present) is assumed to be day 11 area.
−
Version 1.1
+
Sometimes when growth is about 0, the outlines on, eg, day7, may have slightly less area than the original
 +
explant, so the outlines might be mis-ordered then. [actually it may read the correct day off the name of
 +
the i2i image now].
  
−
Usage: correlation [options] im1.i2i im2.i2i out.i2i
+
  note: If only 2 areas (ie, two .roi files) are present in the zip file, they are assumed to be for
−
options:
+
the explant and day7 (not day11).
−
  -d xdim ydim: output dimensions of new image
 
−
(default = 512,512).
 
−
  -S im1scale im2scale: scale intensities with these scale factors. newval = (intens-min_intens)*scale
 
−
  -B b1 b2:    force min_intens (black level) to specified value.  newval1 = (intens-b1)*scale1
 
−
  -i val1 val2: Ignore any voxels in image1 <= val1 and
 
−
any voxels in image2 <= val2.
 
  
−
  -h val1 val2: Ignore any voxels in image1 >= val1 and
+
  note: This program assumes the center of the combined image (usually 4 well images are combined into one) is the
−
any voxels in image2 >= val2.
+
center of the well. see combine_images.pl. It also assumes the combined image name ends in combined.i2i
 +
(see -combined).
  
−
  -r: instead of intensities, use the rank
+
  note: some of the scripts I use to then analyze these results will multiply the area by 2, to get a "true" estimate
−
order of the intensities.
+
  of growth; since the value produced by mask_gedmat_images3.pl will be (roughly) half of the real growth rates.
−
  -a: automatically calculate the size of the
 
−
output image. attempts to make the scale
 
−
factor = 1.
 
−
  -b: keep the xdim and ydim scale factors the same.
 
−
  -s: calculate Spearman rank-order correlation
 
−
coefficients. print results to stdout.
 
−
  -z zslice:    only examine the specified z slice (the first
 
−
slice is considered slice 0).
 
−
note: entire image name is NOT required.
 
−
note: a - in place of an image name means stdin or stdout.
 
  
−
examples:
+
  usage:
−
    # to apply this to a set of 3D images, using the same scales and size images for easy comparison
+
mask_gedmat_images3.pl [options]
−
correlationL -S s1 s2 -d 512 512 -B 0 0 image1.i2i corr1.i2i    (and same for other images)
 
−
 
−
===  create_imagesL ===
 
  
−
This program takes output from many runs of "overlap" and
+
  options:
−
produces two images. One image is a percent colocalization image
+
-tmpdir=fullpathname directory to use for temporary files which are created, eg: -tmpdir=/home/lml/krypton
−
and the other is a probability image.  The overlap program MUST be
 
−
run via the "many_overlaps" command.  The probability calculations
 
−
are the same as those performed by "probcolocal", but only -n
 
−
needs to be specified since the other values are automatically
 
−
extracted from the file produced by "many_overlaps".
 
  
−
The percent image is the percent of voxels above threshold in image1
+
-zip_file=file.zip
−
(as originally specified to "many_overlaps") which overlap with
+
-zip_file=file.zip:file2.zip:...:filen.zip
−
voxels above threshold in image2. A 100 in the output image = 100 percent.
+
only examine the one specified zip file, not all those in the directory
−
The probability of observing as much (or more) overlap as we did due to  
+
(If the second form is used, you can specify as many as you want.)
−
random chance is the prob.i2i image.  Probabilities are mulitplied by 100.
+
note: you may still need to specify -zip_ending so that the ending of this file can be stripped
 +
      off correctly (so the -combined_ending and -results_ending can then be added correctly).
 +
-zip_ending=stuff e.g., -zipending=_area.zip  . All files which end in this will be analyzed; default = _Area.zip
 +
-skip=file.zip[:file2.zip...]  list of zip files (without the full path) to skip.
  
−
Position along the y-axis (vertical axis) of the image, corresponds to
+
-results_ending=stuff e.g., -results_ending=_results.txt .  This is what the -zip_ending should be replaced by
−
thresholds for image2. The x-axis represents image1 thresholds.  The
+
to get the name of the associated fiji results file. note the _ should be included here
−
lowest threshold is the bottom left corner.  See the first line in
+
if it was included in -zip_ending.
−
your datafile for the range of thresholds this represents.
 
−
Overlap produces statistics for a range of allowed overlaps.
 
−
Create_images, by default, chooses an overlap within 1 voxel.
 
−
If you want to use a different overlap distance use the -o option.
 
  
−
See also: overlap, probcolocal, many_overlaps
+
-combined_ending=stuff  e.g., -combined_ending=_combined_reduced_.i2i  . default = combined.i2i.
 +
This is any string which helps identify the last part of the combined image filename.
 +
-combined_scaling=#:# e.g., -combined_scaling=2:2 if reduceima -bin 2 2 was applied to get _combined_reduced_.i2i.
  
−
Usage:
+
-scale=#:# distance and area (eg, -scale=2:4 if reduceima -bin 2 2 was used) scaling. This will multiply
−
create_images [options] datafile percent.i2i prob.i2i
+
the area stored in the fiji file and the area calculated via my mask by the second number.
−
options:
+
The first number will multiply the distance of the explant from the center of the image.
−
  -o distance:    voxels within this distance count as overlapped.
+
This option is just here to handle 001_BAP and 002_PRO datasets. They were analyzed in fiji
−
  -n number:    number of voxels (1000000 default), see probcolocal.
+
with the images reduced by a factor of 2 in x and y. So the fiji areas produced will be low
 +
by 4x, as will my areas.  This option compensates for that.  See ~/Corvera/Raz/README for 10/11/13.  
  
−
Examples:
+
-check_intersect check if outlines intersect with the explant's outline. If so, set an error flag on stdout printout
−
  many_overlaps -t 100 1000 100 -T 100 1000 100 alpha_T.i2i beta_T.i2i tmp_output
+
and print info to stderr. eventually will just use area numbers rather than masking area in this situation.  
−
  create_images -n 363264 tmp_output percent.i2i prob.i2i
+
that is not done yet ...  
−
 
−
== Analysis of Points ==
 
−
===  components.pl  ===
 
  
−
Calculates the number of connected components resolved over a range of point densities.
 
  
−
components.pl [options]
+
-save_day[=directory]
 +
saves the final binary mask for each days growth (eg, day7 growth minus the explant and minus the part growing towards the wall)
 +
into a 3d mask image (patientid_tissue_day#_mask.i2i).  This image will be reduced down in xdim and ydim from the
 +
original.  patientid_tissue_day#_.play_labels will also be created (labels which well each z slice comes from).
 +
This is mostly for quality control purposes.
 +
In addition, patientid_tissue_day#.rpts will be created (similarly reduced down in size) (coords may be off by 1?).
 +
View the file via: addlines maskim.i2i maskim.rpts -  |play.FC14 -P -1 R -labels maskim.play_labels -
 +
  or via: play_results3.pl  
 +
patientid_tissue_day#_orig.i2i will be the original images (also reduced in xdim and ydim, and only one 2d slice
 +
from each original 3d image).
 +
note: older versions of play may need the comment line at the top of maskim.play_labels removed.
 +
note: if you want images to be in another directory, specify a full pathname, eg -save_day=/home/lml/krypton (without last /).
 +
note: any existing file with these names will be OVER-WRITTEN.
 +
note: this option creates lots of temporary files (in tmpdir) of the form __*.i2i, any existing
 +
      files that match this will be DELETED.
 +
see also: montage
  
−
options:
+
-patient5 patient5 files have some odd naming. use this option if analyzing them.
  
−
-numpts=low:hi:delta
+
-use_explant_outline legacy. specify this if you want the center of the explant to be based on the average of the pixel positions
−
-repeat=# number of times to repeat at each point density
+
of just the pixels along the boundary (rather than the center of mass).
−
-res=# in microns
+
-use_bbox_area legacy. specify this if you want to order the outlines based on their bounding box area rather than their true area.
−
-outfile=name name of output file. otherwise to _tmp. always need write permission in current directory.
+
You should probably specify both options (if you want either one), I'm not sure about consistency if only one is specified.
  
−
   
+
-just_check just check whether each zip roi outline has the correct number of associated image files.
−
=== convex_hull.pl  ===
+
eg, if the zip file has 3 outlines (explant, firstday, secondday) there should be two i2i files with DAY# in their name.
 +
prints to stderr info about those which don't have the correct number. does nothing else.
 +
-just_stats goes a bit further than -just_check (does that too), sets -verbose and prints out stats about each outline. then exits.
 +
-verbose some extra info to stdout
 +
-debug prints some tracing info to stderr
 +
-print_fname print name of each zip file as it is being analyzed, to stderr.
 +
-dlevel=# extra debugging. the bigger the number the more the printout. # is an integer starting at 1.
 +
-dlevel=1 will print a trace of commands to execute to mimic (parts of) this perl script.
 +
-noclean don't remove the temporary files which were created
 +
the files are: _ones.i2i _explant.rpts _day11.rpts _day7.rpts _excluded_region.rpts
 +
      _explant.i2i _day11.i2i  _day7.i2i _day7_noex.i2i _day11_noex.i2i _day7_noex_halfex.i2i _day11_noex_halfex.i2i
 +
  (if multiple zip files were analyzed, only the files from the last one will be left, since they get reused)
 +
-dont_recalc only for diagnostics. does NOT restrict the analysis to just the growth away from the wall.
 +
with this specified this entire program should just reproduce the numbers we already
 +
have in *Results.txt since that uses the entire area. ie, this make the program a no-op.
 +
 
 +
  see also:
 +
/home/lml/krypton/Corvera/Raz/README
 +
analyze_gedmat3.pl a script which calls mask_gedmat_images3.pl for all of Raz's data. Also versions 4,5,6, and 7.
 +
play_results3.pl
 +
raz_rates.pl
 +
mask_gedmat_images.pl
 +
mask_gedmat_images2.pl
 +
/storage/big1/raz/GEDMAT/
 +
ReadROI.pl
 +
extract_area.pl
 +
combine_images.pl
 +
gedmat_to_xy.pl
 +
/home/lml/krypton/Corvera/Raz/analyze_gedmat.pl
 +
chres_rpts.pl
 +
  filter_rptsinz.pl
 +
 
 +
  examples:
 +
cd /storage/big1/raz/GEDMAT/006_JTH/OM
 +
 
 +
# generate 006_JTH_OM.areas with all the data (area_mex_border is the number which should replace farea)
 +
# also produce 006_JTH_OM_7masks.i2i and 006_JTH_OM_11masks.i2i so I can check on the validity of the algorithm.
 +
# place diagnostic mask images in /home/lml/krypton/Corvera/Raz/output and the output data file in
 +
# /home/lml/krypton/Corvera/Raz (note: all directories must already exist).
 +
mask_gedmat_images.pl -tmpdir=/home/lml/krypton/Corvera/Raz/output  -zip_ending=_Area.zip -results_ending=_Results.txt
 +
-combined_ending=combined.i2i -days=7:11
 +
-save_day=/home/lml/krypton/Corvera/Raz/output
 +
>& /home/lml/krypton/Corvera/Raz/006_JTH_OM.areas
 +
 
 +
# look at the masks produced:
 +
cd /home/lml/krypton/Corvera/Raz/output
 +
      play_results3.pl 006_JTH_OM_day7
 +
or:
 +
fgrep -v # 006_JTH_OM_11masks.play_labels > tmp.play_labels  # eliminate the comment line, play doesn't like those.
 +
play -labels tmp.play_labels 006_JTH_OM_7masks.i2i
 +
 
 +
# compare the full areas produced which what fiji did (as a check). only pull out the good data. they should be very similar:
 +
cd /home/lml/krypton/Corvera/Raz
 +
gedmat_to_xy.pl -cond='if (($date eq "date_sure")&&($perr==0)&&($aerr==0)) {1;} else {0;}'
 +
-xcoord='$area' -ycoord='$farea' 006_JTH_OM.areas | xmgrace -free -pipe
 +
 
 +
# now grab the data we really want. compare the old way, with the area when the growth half near the well border is masked off:
 +
gedmat_to_xy.pl -cond='if (($date eq "date_sure")&&($perr==0)&&($aerr==0)) {1;} else {0;}'  
 +
-xcoord='$area_mex' -ycoord='$area_mex_border' 006_JTH_OM.areas | xmgrace -free -pipe
 +
 
 +
# it may be hard to see some of these images as they are very large, do it this way (note the -L to keep the -1 boundary values):
 +
addlines _ones.i2i _excluded_region.rpts - | reduceima -bin 3 3 -L - -|play -P -1 R -P -2 G -P -3 B -P -4 C -P -5 Y -
  
−
This program takes an object outputfile (unless -rptsin or -tripletsin) produced by countobjs (or maskobjs) and finds the minimal
 
−
spanning tree connecting the center of mass of the objects, and the convex hull. Writes out connections as an rpts file to stdout.
 
−
It also loops through 20 thresholds  for the pts, and prints to stderr some stats about the minimal spanning tree and
 
−
convex hull of the points above each of these thresholds.
 
  
−
Options can be used to only selected a subset of the objects and to constrain the maximum distance (-maxdist).  
+
# run with some diagnostics:
 +
cd /storage/big1/raz/GEDMAT/006_JTH/OM
 +
/home/lml/krypton/bin/mask_gedmat_images.pl -tmpdir=/home/lml/krypton/Corvera/Raz -zip_ending=_Area.zip
 +
        -results_ending=_Results.txt -combined_ending=combined.i2i -days=7:11
 +
        -save_day=/home/lml/krypton/Corvera/Raz/006_JTH_OM_7masks.i2i    > /home/lml/krypton/Corvera/Raz/006_JTH_OM.areas
 +
# play.FC14 allows labels to have comment lines (first character of line is a #)
 +
cd home/lml/krypton/Corvera/Raz
 +
play.FC14 -labels 006_JTH_OM_11masks.play_labels 006_JTH_OM_11masks.i2i
 +
chres_rpts.pl -sx=.33 -sy=.33 -truncate -min=1:1 output/_day11.rpts > _day11_bin3.rpts
 +
chres_rpts.pl -sx=.33 -sy=.33 -truncate -min=1:1 output/_day7.rpts > _day7_bin3.rpts
 +
chres_rpts.pl -sx=.33 -sy=.33 -truncate -min=1:1 output/_explant.rpts > _explant_bin3.rpts
 +
reduceima -bin 3 3 output/_day11_noex_halfex.i2i - | addlines - _day11_bin3.rpts - | addlines - _day7_bin3.rpts - |
 +
addlines - _explant_bin3.rpts - | play -P -1 R -
 +
addlines /storage/big1/raz/GEDMAT/006_JTH/OM/D07_20130524_GEDMAT_006_JTH_OM_DAY7_combined.i2i output/_day11.rpts - |
 +
addlines - output/_day7.rpts - | addlines - output/_explant.rpts - | reduceima -bin 3 3 -L - - | play -P -1 R -
  
−
It was written with the hope that triplets (maxima in an image which has a cell in the center) will show a pattern in their
 
−
convex hull area, so that when the threshold rises high enough, only maxima inside the cell will still be present and hence
 
−
the convex hull will be stable and of much smaller area.
 
  
−
This program requires perl modules Graph::Kruskal and Math::Geometry::Planar, use cpanp to install these:
+
# do a bunch of stuff
−
    yum install perl-CPANPLUS-0.91.3-202.fc16.noarch   - for cpanp
+
analyze_gedmat3.pl # apply this perl script to lots of data
−
    cpanp -i Graph::Kruskal
+
cd output
−
    cpanp i Math::Geometry::Planar
+
      play_results3.pl 006_JTH_OM_day7   # look at some of it
 +
raz_rates.pl # generate stats
 +
  
−
usage:
 
−
  convex_hull.pl -tripletsin file1.triplets  > file1_mst.rpts
 
  
−
   input options:
+
   source code: in /home/lml/krypton/bin/mask_gedmat_images.pl
−
    -dx=#   spacing in the x direction, default = 1
 
−
    -dy=#   spacing in the y direction, default = 1
 
−
    -dz=#   spacing in the z direction, default = 1
 
−
    -rptsin     instead of file1.objs, read file1.rpts where file1.rpts was produced by max2d.
 
−
      -size1 and -iod1 are ignored when this option is specified.
 
−
    -tripletsin   instead of file1.objs read in file1.xyi,  triplets of x y and intensity.
 
  
−
  analysis options:
+
−
    -size1=min:max     min and max object size allowed for file1.objs in pixels (inclusive)
+
</nowiki>
−
    -iod1=min:max   min and max object iod allowed for file1.objs (inclusive), or intensity if -tripletsin was specified.
+
===  membrane3D.pl  ===
−
    -maxdist=#: :  don't allow connections between pts further apart than this distance (after applying dx, dy, dz spacing factors).
+
<nowiki>
−
  warning: if the distance is too small, then all the pts will not be connected to begin with;
+
 
−
    this may cause the minimal spanning tree algorithm to fail.
+
A script to run the routines necessary to perform 3D membrane segmentation.
−
    default maxdist is  3000
+
note: it needs write permission in the directory where inputimage.i2i resides since all output images go there too.  
 +
It will run the following programs, in order:
 +
  1. /home/lml/krypton/facil/i2i2mhd (convert to mhd format)
 +
  2. /home/lml/krypton/segmentation/MR-Source/ACME/ACME-Binary/bin/cellPreprocess (median filter, fill holes)
 +
  3. /home/lml/krypton/segmentation/MR-Source/ACME/ACME-Binary/bin/resample (resample pixel sizes)
 +
  4. /home/lml/krypton/segmentation/MR-Source/ACME/ACME-Binary/bin/multiscalePlateMeasureImageFilter  (determine the planarity of each region)
 +
  5. /home/lml/krypton/segmentation/MR-Source/ACME/ACME-Binary/bin/membraneVotingField3D (update planarity based on neighbors voting)
 +
  6. /home/lml/krypton/segmentation/MR-Source/ACME/ACME-Binary/bin/membraneSegmentation  (produce segmented regions via a watershed algorithm)
 +
Each program expects input files of a certain type (e.g. unsigned char).
  
−
  output options:
 
−
    -addlines :  output rpts format which addlines is more likely to be happy with (may not work in dave?)
 
−
    -hist :  calculate histogram of distances (hist[int(dist)]++, where int just truncates to integer). obsolete.
 
−
    -sizehist :  calculate histogram of object sizes (in voxels) in file1.out. (hist[int(size)]++, where int just
 
−
        truncates to integer). If you don't care about distances between two files (ie, only want -sizehist)
 
−
        put the same file name for both files on the command line.  obsolete.
 
−
    -rpts=file.rpts :  put bounding boxes around objects found and lines between closest objects (similar to objs2bb.pl)
 
−
  display this in dave eg:  dave -n 0 -u -I file.i2i file.rpts (use the -n 0 and -u options)
 
−
  note: may not show up in dave until you click on "scale" to force a rescale (or draw?).
 
−
    -jack=max.rpts:# :  create an rpts file with the maxima in each object displayed as a small jack.
 
−
    # controls the size of the jack. 1=1 pixel in size, 2 = 2 pixels, etc.
 
−
-twod   add this (in addition to -jack) if you want 2D jacks rather than 3D jacks.
 
−
    -strlim=# :  limit length of names of objects in rpts files to # characters.  dave sometimes crashes if the
 
−
        names are too long.
 
−
    -tyz :  transpose coords in yz before writing them to the rpts (must specify -rpts option)
 
−
    -color=# :  for the w value (color) use the specified value (use -P in play to see this as a color). default = none.
 
−
        if you have no color (no w value), addlines can put one in (-P option in addlines).
 
  
−
  diagnostics:
+
usage:
−
    -silent :  suppress warning messages.
+
  membrane3D.pl [options] inputimage.[i2i|mhd|mha] outimage.mhd
−
    -verbose
 
−
-debug
 
  
−
example:
+
output files:
−
   convex_hull2.pl -tripletsin -jack=jack.rpts:5 -addlines -twod -maxdist=100 file1.triplets  > file1_mst.rpts
+
   inputimage.mhd produced by i2i2mhd  - short ints
−
   addlines2 -P -1 image.i2i file1_mst.rpts - | addlines2 -P -2 - jack.rpts - | reduceima -bin 3 3 -L - -| play -P -1 R -P -1 G -
+
  inputimage_med.mhd produced by cellPreprocess
 +
  inputimage_med_resamp.mhd produced by resample, unsigned char
 +
  inputimage_med_resamp_planar.mhd produced by multiscalePlateMeasureImageFilter, floating pt (doubles) between 0 and 1, how "planar" a pt is.
 +
   inputimage_med_resamp_eigen.mhd produced by multiscalePlateMeasureImageFilter, an eigenmatrix at each pt
 +
  inputimage_med_resamp_eigen_voting.mhd produced by membraneVotingField3D, result of voting, unsigned char
 +
  outimage.mhd produced by membraneSegmentation, result of watershed, each region labeled with an id (short integer).
  
 +
note: all units below are in the same units as the pixel units (I think). So if pixel spacing is 2 (um) in x, and a radius
 +
      of 2 is specified, that is a one pixel radius.
  
−
see also:
+
step 1. options for converting i2i to mhd:
−
   convex_hull, objs2mst.pl
+
  -convert=dx:dy:dz   The first step is converting our i2i format to mhd format. This specifies the pixel spacing
−
   objs2bb.pl, objs2dist.pl, objs2dist121.pl, closest_object, closest_voxel, countobjs, maskobjs, objs_via_ascent
+
in x y and z directions for that. default assumes 1:1:1. The "conversion" really just creates inputimage.mhd which
−
  resolve_pts
+
is a text header which points to the original i2i image and gives some info about it.    
 +
If the input image has a .mhd or .mha extension already (instead of .i2i) then no conversion is done.
 +
Olga's images acquired with a 10x objective have a pixel size of .645 um/pixel (and z spacing of 10 um).
 +
Example: G03_x_1_1_binned.i2i has a pixel size of 2.5x2.5x10.  So -convert=2.5:2.5:10 specifies this in the mhd header.
  
−
+
step 2. options for cellPreproces:
−
===  objs2alldists.pl  ===
+
this may produce an unsigned char image (?)
−
Like obs2disp.pl but calculates the distance to ALL the objects (within -maxdist), not just the closest.
+
   -median=# radius of median filtering preprocessing (and hole filling). If this is not specified it isn't done.
−
See ~/krypton/Zhuge/grants/AsthmaRO1/README for 5/27/11 and /home/lml/krypton/Zhuge/BKandRyr/analysis6/run18
 
−
   (which did NOT use this).
 
  
−
This program takes 2 object outputfiles produced by countobjs (or maskobjs) and finds the shortest distance
+
step 3. options for resampling:
−
from the center of mass of objects in file1.out to the center of mass of objects in file2.out.
+
this may produce an unsigned char image (?)
 +
  -resample=x:y:z  resample in the x y and z directions, eg. -resample=2:2:1 will halve the x and y dimensions and leave z unchanged.
 +
note: this will change the spacing between the pixels; but the routines which follow will understand that.
 +
For example: if after resampling, the pixels are spaced 10 um apart, then -planar=10 will use a sigma of 1 pixel.
 +
If this is not specified it isn't done.
  
−
If both file arguments are the same (e.g., objs2dist.pl file1.out file1.out >file.dist), you can use this
+
  note: it may be necessary to do the above operations, which also convert the image into unsigned char, which is required for
−
routine to find the distances between objects within an image (an object match with the same id is automatically
+
step 4, multiscalePlateMeasureImageFilter?
−
ignored in this case so objects don't match to themselves). See also the option -skipsame.
+
  note: step 6 (watershed) may also work best if the pixel sizes are the same in all directions.
  
−
(modified from /storage/big1/lml/was_voxel/Corvera/track/objs2bb.pl)
+
step 4. options relating to planarity:
 +
the planarity image produced will be doubles from 0-1.
 +
  -planar=# the sigma of the Gaussian used when classifying pixels as planar or not. default = 1
 +
  -alpha=# see the help for /home/lml/krypton/segmentation/MR-Source/ACME/ACME-Binary/bin/multiscalePlateMeasureImageFilter
 +
  -beta=# ditto
 +
  -gamma=# ditto
 +
  -c=# ditto
 +
  -planaropts="options for multiscalePlateMeasureImageFilter here" 
 +
available options (so far, but anything specified here will be passed), : -a # -b # -g # -c # -d -s -B -T
 +
options which go here should be specified in the format that multiscalePlateMeasureImageFilter wants them,
 +
for example: -planaropts="-c 5 -d -B"
 +
not in the alternative format that Membrane3D.pl desires (e.g. -planaropts="-a 5" is correct, -planaropts="-alpha=5" is wrong).
 +
    -stop_after_planar don't do anything after this step (e.g. voting for planar patches may not make sense if -B or -T specified)
  
−
usage:
+
step 5. options relating to voting:
−
  objs2alldists.pl file1.out file2.out >file.dist 
+
the resulting tensor vote image will be unsigned chars from 0-255.
 +
  -region=# the sigma which defines the region within which a pixel influences other pixels. default = 10
  
−
options:
+
step 6. options relating to watershed segmentation:
−
  -dx=# :  spacing in the x direction ,default = 1
+
The resulting image will be long ints (id numbers) of regions.  
−
  -dy=# : spacing in the y direction ,default = 1
+
                See the help for /home/lml/krypton/segmentation/MR-Source/ACME/ACME-Binary/bin/membraneSegmentation
−
  -dz=# :  spacing in the z direction ,default = 1
+
  -thresh=# threshold value. this is applied to the tensor voting image to turn it into a binary image. then a distance
−
  -size1=min:max :  min and max object size allowed for file 1 in pixels (inclusive)
+
transform and watershed are applied. default = 128
−
  -size2=min:max :  min and max object size allowed for file 2 in pixels (inclusive)
+
  -segopts="options for membraneSegmentation here"   
−
  -iod1=min:max :  min and max object iod allowed for file 1 (inclusive)
+
for example:  -segopts="-S -l 20 -a 5 -c -D" will use this exact string as options.
−
  -iod2=min:max :  min and max object iod allowed for file 2 (inclusive)
+
options so far: -S -l # -a # -s # -n -i # -w # -m # -c -v -d -D
−
  -maxdist=#: :  don't allow matches further away than this distance (after applying dx, dy, dz spacing factors).
 
−
  default maxdist is ~  3000.
 
−
  -hist :  calculate histogram of distances (hist[int(dist)]++, where int just truncates to integer).
 
−
  -sizehist : calculate histogram of object sizes (in voxels) in file1.out. (hist[int(size)]++, where int just
 
−
  truncates to integer). If you don't care about distances between two files (ie, only want -sizehist)
 
−
  put the same file name for both files on the command line.
 
−
  -rpts=file.rpts :  put bounding boxes around objects found and lines between closest objects (similar to objs2bb.pl)
 
−
  display this in dave eg:  dave -n 0 -u -I file.i2i file.rpts (use the -n 0 and -u options)
 
−
  note: may not show up in dave until you click on "scale" to force a rescale (or draw?).
 
−
  -jack=max.rpts:# :  create an rpts file with the maxima in each object displayed as a small jack.
 
−
  # controls the size of the jack. 1=1 pixel in size, 2 = 2 pixels, etc.
 
−
  -strlim=# :  limit length of names of objects in rpts files to # characters.  dave sometimes crashes if the
 
−
  names are too long.
 
−
  -tyz :  transpose coords in yz before writing them to the rpts (must specify -rpts option)
 
−
  -skipsame :  don't match an object in file1.out to an object with the same id in file2.out
 
−
  This is set automatically if file1.out = file2.out.
 
−
  -silent :  don't print the distances (sets -hist)
 
−
  -simpleprint :  when printing distances, just print distance, one per line, nothing else. to be read in
 
−
  by ~/Zhuge/grants/AsthmaRO1/distance_distribution.m. note: that program assumes distances
 
−
  are in pixels and pixels are 80 nm, unless you specify pixelsize=somethingelse.
 
−
  -simpleprint2 :  like simpleprint, but the first column is id of the Ryr to which this BK is closest.
 
−
  -onelineprint :  like objs2alldists.pl  but prints the distance to ALL the objects (within -maxdist),
 
−
  not just the closest on one line, so we can keep track of which BK are near which Ryr.
 
−
  see comdists3.pl (which uses this option).
 
−
  -verbose
 
  
 +
other options:
 +
  -verbose echo what is being done to stderr
 +
  -dummy don't really do anything; echo what would have been done to stderr.
 +
 
 
example:
 
example:
−
  countobjs -k -t 2000 2000 wave1.objs wave1.i2i
+
reduceima -bin 4 4 G03_x_1_1.i2i G03_x_1_1_binned.i2i    - go from .645um/pixel, 10um in z -> 2.5um/pixel, 10um in z
−
  countobjs -k -t 2000 2000 wave2.objs wave2.i2i
+
# -convert: just put correct pixel size info into an mhd header
−
  objs2dist.pl wave1.objs wave2.objs > wave1and2.dist
+
# -median:  smooth with a filter 10 um (4 pixels, since they are now 2.5um/pixel) in radius (diameter?)
 +
# -resample: change to a square pixel (10 um in x y and z). Change image format to unsigned char (needed by MultiscalePlateMeasureImageFilter)
 +
# -planaropts: options for planarity filter
 +
# -region:  width (in um) of neighbors for voting on planarity and updating it.
 +
# -segopts:
 +
# -thresh:  threshold for watershed segmentation
 +
membrane3D.pl -convert=2.5:2.5:10 -median=10 -resample=2:2:1 -planaropts="  " -region=50 -segopts="  "  -thresh
 +
G03_x_1_1_binned.i2i G03_x_1_1_results.mhd
  
−
see also:
+
see also: /home/lml/krypton/segmentation/MR-Source/ACME/lmltest/README
−
  objs2dist121.pl, closest_object, closest_voxel, countobjs, maskobjs, objs_via_ascent, pt_distances
+
  /home/lml/krypton/segmentation/FindingCellMembranesIn3D.pdf
 +
          i2i2mhd - convert i2i into mhd format
 +
  mhd2i2i - convert an mhd image (but not an eigenimage)
 +
  RegionId2Mesh - will convert the ID image produced into a mesh image or rpts file
 +
  mayavi2  - to visualize *.mha and *.mhd files.
 +
  Fiji - my version has a File-> Import metaimage  plugin
  
 
   
 
   
−
===  objs2dist2.pl  ===
+
</nowiki>
−
objs2dist2.pl: like objs2dist.pl but when -rpts specified prints out lines to ALL objects
+
===  Norio_stats.pl  ===
−
      within -maxdist (which now defaults to 20), rather than just the closest. 
+
<nowiki>
−
      So we can see all the BK clusters which are close enough
+
 
−
      to a Ryr to be activated. see /home/lml/krypton/Zhuge/paper2010/README for 5/20/10.
+
This programs takes each pts file in the current directory, thresholds the associated vinculin image,
 +
applies that mask to the trip6 image, and produces a .coloc file from the masked trip6 image (using mask_and_coloc.pl);
 +
overlap statistics are then calculated (and put into a file name *_masked.coloc where * is the root of the trip6 image name)
 +
and finally the highest ratio data from each image pair is printed to stdout (using sort_overlap3.pl).
  
−
This program takes 2 object outputfiles produced by countobjs (or maskobjs) and finds the shortest distance
+
stats.pl [options]  > stats.out
−
from the center of mass of objects in file1.out to the center of mass of objects in file2.out.
+
  -nothing : do nothing, just echo what would be done
 +
  -h : print this help
 +
  -thresh=#    : threshold for the vinculin image. default = 20
 +
  -regions=#:#  : only allow mask regions within the specified min:max size (in pixels, inclusive).
 +
: Also creates *_count.i2i  and *.countobjs (* = root of vinculin image name) intermediate files.
 +
: note: these counts are for the vinculin image after the mask (pts file) is applied.
 +
  -clean:      : clean up (ie, remove) *_count.i2i and *.countobjs images (if -regions specified)
  
−
If both file arguments are the same (e.g., objs2dist.pl file1.out file1.out >file.dist), you can use this
+
−
routine to find the distances between objects within an image (an object match with the same id is automatically
+
</nowiki>
−
ignored in this case so objects don't match to themselves). See also the option -skipsame.
+
===  run2_Ronghua.pl ===
 +
<nowiki>
  
−
(modified from /storage/big1/lml/was_voxel/Corvera/track/objs2bb.pl)
+
this program goes through Ronghua's directory (see -dir option)
 +
analyzing files.  output is put into the current working directory (ie, the directiory
 +
run.pl is run from).
 +
A tag in the name of the form Ryr_thresh_minsize_maxsize_Bk_thresh_minsize_maxsize
 +
is added to the output name  of name.patchdist  (nearest voxel distance between whole
 +
patches) and name.comdist (distances between nearest center of mass of the patches).
  
 
usage:
 
usage:
−
   objs2dist2.pl file1.out file2.out >file.dist 
+
   run.pl [options]
  
 
options:
 
options:
−
   -dx=# : spacing in the x direction ,default = 1
+
   -dir=directorypath    : without the final slash. directory to analyze. default = /mnt/big3/rzg/mtc/immunocytochemistry/july05/imcc/70105/epr
−
   -dy=# : spacing in the y direction ,default = 1
+
   -ryrthresh=# : threshold for ryr (488) images, default =  
−
   -dz=# : spacing in the z direction ,default = 1
+
   -min_ryrsize=# : minimum allowed object size in pixels, default = 5
−
   -size1=min:max :  min and max object size allowed for file 1 in pixels (inclusive)
+
   -max_ryrsize=# : max allowed object size in pixels, default = 150
−
   -size2=min:max :  min and max object size allowed for file 2 in pixels (inclusive)
+
   -min_ryriod=# : minimum allowed object iod, default = 354
−
   -iod1=min:max : min and max object iod allowed for file 1 (inclusive)
+
   -max_ryriod=# : maximum allowed object iod, default = 10000000
−
   -iod2=min:max : min and max object iod allowed for file 2 (inclusive)
+
 
−
   -maxdist=#: : don't allow matches further away than this distance (after applying dx, dy, dz spacing factors).
+
   -bkthresh=# : threshold for bk (594) images, default =
−
  default maxdist is 20.
+
   -min_bksize=# : minimum allowed object size in pixels, default = 5
−
   -hist : calculate histogram of distances (hist[int(dist)]++, where int just truncates to integer).
+
   -max_bksize=# : max allowed object size in pixels, default = 50
−
   -sizehist : calculate histogram of object sizes (in voxels) in file1.out. (hist[int(size)]++, where int just
+
   -min_bkiod=# : minimum allowed object iod, default = 4686
−
  truncates to integer). If you don't care about distances between two files (ie, only want -sizehist)
+
   -max_bkiod=# : maximum allowed object iod, default = 10000000
−
  put the same file name for both files on the command line.
+
 
−
   -rpts=file.rpts : put bounding boxes around objects found and lines from objects in file1.out to all objects
+
   -dx=# : relative distance in x, default = 1  
−
  in file2.out that are <= maxdist away (these lines will become an object named "near_to_file1.out").
+
   -dy=# : relative distance in y, default = 3
−
  display this in dave eg:  dave -n 0 -u -I file.i2i file.rpts (use the -n 0 and -u options)
+
   -dz=# : relative distance in z, default = 1
−
  note: may not show up in dave until you click on "scale" to force a rescale (or draw?).
+
 
−
   -jack=max.rpts:# : create an rpts file with the maxima in each object displayed as a small jack.
+
   -noclean
−
  # controls the size of the jack. 1=1 pixel in size, 2 = 2 pixels, etc.
+
   -debug
−
   -strlim=# : limit length of names of objects in rpts files to # characters.  dave sometimes crashes if the
 
−
  names are too long.
 
−
   -tyz : transpose coords in yz before writing them to the rpts (must specify -rpts option)
 
−
   -skipsame :  don't match an object in file1.out to an object with the same id in file2.out
 
−
  This is set automatically if file1.out = file2.out.
 
−
   -silent :  don't print the shortest distances (sets -hist)
 
 
   -verbose
 
   -verbose
  
−
example:
+
−
   countobjs -k -t 2000 2000 wave1.objs wave1.i2i
+
</nowiki>
−
   countobjs -k -t 2000 2000 wave2.objs wave2.i2i
+
===  countobjs  ===
−
   objs2dist2.pl wave1.objs wave2.objs > wave1and2.dist
+
<nowiki>
 +
Usage:
 +
   countobjs [options] outputfile image1.i2i [image2.i2i]
 +
Description:
 +
  Count objects in co-localized images. If only one image given
 +
  the objects will be counted in that one image.  Copious amounts
 +
  of information will be written to the output file.
 +
  A - for outputfile will write to stdout.
 +
  outpufile can be analyzed with objs2bb.pl and objs2dist.pl
 +
 
 +
Options:
 +
  -C # #        : set opacity and brightness level to these values (0-255 only)
 +
  -k             : use original images for data analysis (so scale and black level not important and
 +
      -t option can be in same units as original, unscaled, image values)
 +
  -S # # #      : image # scale and black level (default autoscale image)
 +
  -r # # # # # # : region of image to count
 +
                  (-r x0 y0 z0    x1 y1 z1)
 +
                  (  ^lower left  ^upper right)
 +
                  (zero indexed coordinates)
 +
                  (default whole image)
 +
  -s # #        : size of smallest and largest objects to count
 +
                  (default 1-100000)
 +
  -m # #        : min and max iod of allowable objects (applied to image1 only).
 +
  -t thresh1 thresh2    : thresholds (applied after -S or default scaling, unless -k specified).
 +
                  Like left brightness slider in DAVE.
 +
                  if only one image, second number still required, but ignored.
 +
  -o <image.i2i> : output image with all objects represented by the voxel
 +
                  with the maximum intensity in the object
 +
                  the intensity of the voxel will be the IOD of the object.
 +
  -O <image.i2i> : output image with all pixels in each object having
 +
                  the intensity of the original objects IOD.
 +
                  NOTE: this image will be in floating point format.
 +
                        use float2int to convert it back to "normal" format.
 +
   -Q <image.i2i> : output image with all pixels in each object having
 +
                  a unique id # (1-# of objects). Must be < 32766 objects.
 +
                  NOTE: this image will be in standard signed short int format.
 +
  -M #          : When countobjs is given 2 images, this flag will
 +
                  turn of some subset of the voxels for colcalization.
 +
                  calculations.
 +
                  0: turn off colcalized voxels (default on)
 +
                  1: turn off image 1 voxels (default on)
 +
                  2: turn off image 2 voxels (default on)
 +
  -q     : quiet. don't print out all objects, but just summary statistics
 +
  -Z            : simple analysis, no sliders, opacity,etc. can use with -t option.
 +
  -h            : show this message.
 +
  -H            : show even more help.
 +
 
 +
Notes:
 +
  A - in place of an image name means stdin or stdout.
 +
  Source code in /storage/big1/lml/jac/was_invitro/Projects/countobjs
 +
 
 +
 
 +
</nowiki>
 +
===  countobjsmax3d  ===
 +
<nowiki>
 +
Usage:
 +
  countobjsmax3d [options] -Q imageid.i2i outputfile image1.i2i
 +
Description:
 +
  This version of countobjs finds 3d maxima in image1.i2i, then finds all pixels connected
 +
  to each maxima and downhill from it.  These are labelled with a unique id number and stored
 +
  in imageid.i2i (specified with the -Q option, which is REQUIRED for this version)
 +
  You may want to display imageid.i2i after using colorize_image on it.
 +
  Not tested yet.
 +
 
 +
   Count objects in co-localized images. If only one image given
 +
  the objects will be counted in that one image.  Copious amounts
 +
  of information will be written to the output file.
 +
  A - for outputfile will write to stdout.
 +
  outpufile can be analyzed with objs2bb.pl and objs2dist.pl
 +
 
 +
Options:
 +
  -r # # # # # # : region of image to count
 +
                  (-r x0 y0 z0    x1 y1 z1)
 +
                  (  ^lower left  ^upper right)
 +
                  (zero indexed coordinates)
 +
                  (default whole image)
 +
  -s # #        : size of smallest and largest objects to count
 +
                  (default 1-100000)
 +
  -m # #        : min and max iod of allowable objects (applied to image1 only).
 +
  -t thresh1 thresh2    : thresholds (applied after -S). Like left brightness slider in DAVE.
 +
                  if only one image, second number still required, but ignored.
 +
  -o <image.i2i> : output image with all objects represented by the voxel
 +
                  with the maximum intensity in the object
 +
                  the intensity of the voxel will be the IOD of the object.
 +
  -O <image.i2i> : output image with all pixels in each object having
 +
                  the intensity of the original objects IOD.
 +
                  NOTE: this image will be in floating point format.
 +
                        use float2int to convert it back to "normal" format.
 +
  -Q <image.i2i> : output image with all pixels in each object having
 +
                  a unique id # (1-# of objects). Must be < 32766 objects.
 +
                  NOTE: this image will be in standard signed short int format.
 +
  -M #          : When countobjs is given 2 images, this flag will
 +
                  turn of some subset of the voxels for colcalization.
 +
                  calculations.
 +
                  0: turn off colcalized voxels (default on)
 +
                  1: turn off image 1 voxels (default on)
 +
                  2: turn off image 2 voxels (default on)
 +
  -q     : quiet. don't print out all objects, but just summary statistics
 +
  -Z            : simple analysis, no sliders, opacity,etc. can use with -t option.
 +
  -h            : show this message.
 +
  -H            : show even more help.
 +
 
 +
Notes:
 +
  A - in place of an image name means stdin or stdout.
 +
  Source code in /storage/big1/lml/jac/was_invitro/Projects/countobjs
 +
 
 +
 
 +
</nowiki>
 +
===  threshold.pl  ===
 +
<nowiki>
 +
threshold.pl -numvox=#  [-produce=newimage.i2i] image.i2i
 +
 
 +
calculate the largest threshold for image.i2i which has at least # voxels
 +
equal or above it.  Prints the threshold and the number above threshold to stdout.
 +
 
 +
options:
  
−
see also:
+
   -produce=newimage.i2i    : take the calculated threshold, apply it to image.i2i to produce newimage.i2i
−
   objs2dist121.pl, objs2dist.pl, closest_object, closest_voxel, countobjs, maskobjs, objs_via_ascent, pt_distances
 
  
 
   
 
   
−
===  objs2mst.pl  ===
+
</nowiki>
 +
===  thresholdz.pl  ===
 +
<nowiki>
 +
thresholdz.pl -numvox=#  [-produce=newimage.i2i] image.i2i
 +
 
 +
calculate the largest threshold for image.i2i which has at least # voxels
 +
equal or above it.  Prints the threshold and the number above threshold to stdout.
 +
 
 +
options:
  
−
This program takes an object outputfiles (unless -rptsin) produced by countobjs (or maskobjs) and finds the minimal
+
  -produce=newimage.i2i    : take the calculated threshold, apply it to image.i2i to produce newimage.i2i
−
spanning tree connecting the center of mass of the objects. Writes out connections as an rpts file to stdout.
 
−
Options can be used to only selected a subset of the objects and to constrain max distance (so multiple
 
−
trees can be produced).
 
  
−
usage:
+
−
  objs2mst.pl file1.objs > file1_mst.rpts
+
</nowiki>
 +
===  2Ddistances  ===
 +
  <nowiki>
  
−
options:
+
Calculates a histogram of the amount of light as a function of distance from boundary (0 valued) voxels as specified in the mask.
−
  -dx=# :  spacing in the x direction, default = 1
+
Applies a 2D mask to a 2D time series.
−
  -dy=# :  spacing in the y direction, default = 1
 
−
  -dz=# :  spacing in the z direction, default = 1
 
−
  -size1=min:max :  min and max object size allowed for file1.objs in pixels (inclusive)
 
−
  -iod1=min:max :  min and max object iod allowed for file1.objs (inclusive)
 
−
  -rptsin :  instead of file1.objs, read file1.rpts where file1.rpts was produced by max2d.
 
−
  -size1 and -iod1 are ignored when this option is specified.
 
−
  -addlines :  output rpts format which addlines is more likely to be happy with (may not work in dave?)
 
−
  -maxdist=#: :  don't allow matches further away than this distance (after applying dx, dy, dz spacing factors).
 
−
  default maxdist is ~  3000.
 
−
  -hist :  calculate histogram of distances (hist[int(dist)]++, where int just truncates to integer).
 
−
  -sizehist :  calculate histogram of object sizes (in voxels) in file1.out. (hist[int(size)]++, where int just
 
−
  truncates to integer). If you don't care about distances between two files (ie, only want -sizehist)
 
−
  put the same file name for both files on the command line.
 
−
  -rpts=file.rpts :  put bounding boxes around objects found and lines between closest objects (similar to objs2bb.pl)
 
−
  display this in dave eg:  dave -n 0 -u -I file.i2i file.rpts (use the -n 0 and -u options)
 
−
  note: may not show up in dave until you click on "scale" to force a rescale (or draw?).
 
−
  -jack=max.rpts:# :  create an rpts file with the maxima in each object displayed as a small jack.
 
−
  # controls the size of the jack. 1=1 pixel in size, 2 = 2 pixels, etc.
 
−
  -strlim=# :  limit length of names of objects in rpts files to # characters.  dave sometimes crashes if the
 
−
  names are too long.
 
−
  -tyz :  transpose coords in yz before writing them to the rpts (must specify -rpts option)
 
−
  -color=# :  for the w value (color) use the specified value (use -P in play to see this as a color). default = none.
 
−
  if you have no color (no w value), addlines can put one in (-P option in addlines).
 
−
  -silent :  don't print the shortest distances (sets -hist)
 
−
  -verbose
 
  
−
example:
+
Usage:
−
  countobjs -k -t 2000 2000 wave1.objs wave1.i2i
+
2Dhistogram [options] inimage.i2i inmask.i2i
−
  objs2mst.pl wave1.objs  > wave1mst.rpts
+
Options:
−
  dave -I wave1.i2i -n 0 -u wave1mst.rpts
+
  -v: verbose
  
−
see also:
+
Source code in ~krypton/lml/facil 
−
   objs2bb.pl, objs2dist.pl, objs2dist121.pl, closest_object, closest_voxel, countobjs, maskobjs, objs_via_ascent
+
   -d dimage.i2i : save the distance image as dimage.i2i (converted to ints by casting)
−
  resolve_pts
+
  -n: don't normalize the histogram by the number of voxels in the mask at that distance
 +
  -a #:  average together this many time pts for output histogram. default = 20
 +
see also: histogramL, histogram2
  
 +
Copyright 2010 University of Massachusetts Medical School and the Biomedical Imaging Group
 +
All rights reserved. Explicit permission to use this program must be received prior to use.
 
   
 
   
−
===  objs_within_dist.pl ===
+
</nowiki>
 +
===  analyze_histo ===
 +
<nowiki>
 +
# /home/lml/krypton/bin/analyze_histo
  
−
This program takes 2 object outputfiles produced by countobjs (or maskobjs) and finds the number of objects
+
Analyzes the output histogram image produced by histogram2 via the -f option.
−
in the second file which are within a given distance of each object in the first file. A histogram is
+
Prints out 2 columns: first column is bin number, second column is value at the cutoff.
−
printed out.
+
This is similar to the program "threshold" but instead of one threshold value for the
−
Distances are from the center of mass of objects in file1.out to the center of mass of objects in file2.out.
+
entire image, it gives one at each distance (from a membrane).
−
(modified from objs2dist.pl)
 
  
−
usage:
+
Usage: analyze_histo [options] aimage_H.i2i
−
  objs_within_dist.pl file1.out file2.out >file.dist 
+
options:
 +
-v: calculate cutoff based on percentage of voxels instead of percentage of light
 +
-f #: fractional cutoff (e.g., .80 = 80%), default = 0.950000
 +
-w #: number of bins for running average (default = 5), should be odd.
 +
-t #: only look at voxels >= intensity (must be >= 0), default = 1
  
−
options:
+
Examples:
−
  -dx=# :  spacing in the x direction ,default = 1
+
histogram2 -f a_H.i2i -O a_H.histo -o -p 0 0 0 -d 40 -z 3 mask.i2i a.i2i b.i2i
−
  -dy=# :  spacing in the y direction ,default = 1
+
analyze_histo -f .95 a_H.i2i > a_H.cutoffs
−
  -dz=# :  spacing in the z direction ,default = 1
+
 
−
  -size1=min:max :  min and max object size allowed for file 1 in pixels (inclusive)
+
See also: histogram, histogram2, threshold
−
  -size2=min:max :  min and max object size allowed for file 2 in pixels (inclusive)
+
 
−
  -iod1=min:max :  min and max object iod allowed for file 1 (inclusive)
+
source code in /storage/big1/lml/jac/was_invitro/Projects/boundary/boundary
−
  -iod2=min:max :  min and max object iod allowed for file 2 (inclusive)
+
   
−
  -maxdist=#: : don't allow matches further away than this distance (after applying dx, dy, dz spacing factors).
+
</nowiki>
−
  default maxdist is ~ 3000.
+
=== analyze_yaodl ===
−
  -rpts : read in file in rpts format
+
  <nowiki>
−
  -verbose : extra info written to stderr
 
−
  -vverbose : print other info. can be combined with -verbose.
 
  
−
example:
+
This program reads in a yaodl file as produced by DAVE (volume widget menu: volume to surface ...)
−
  countobjs -k -t 2000 2000 wave1.objs wave1.i2i
+
(hmc produces an ascii format which analyze_yaodl can't handle).
−
  countobjs -k -t 2000 2000 wave2.objs wave2.i2i
+
It identifies connected surfaces (since DAVE considers all surfaces from one pass as one object).
−
  objs_within_dist.pl wave1.objs wave2.objs > wave1and2.dist
+
It then calculates and prints the volume and surface area of each connected surface. Optionally
 +
it will write out a yaodl files (-y) for the largest surface found for each object, with better normals.
 +
This surface can be read back into DAVE with -y option.
  
−
see also:
+
Usage: analyze_yaodl [options] infile.ydl
−
  objs2dist121.pl, closest_object, closest_voxel, countobjs, maskobjs, objs_via_ascent, pt_distances
+
options:
 +
  -y largest.ydl:  write out largest surface of eac object to largest.ydl
 +
  -i image.i2i:  read in the image file associated with infile.ydl (not needed for anything yet).
 +
        -n: recalculate "better" normals (use this with the -y option)
 +
(dave used to produce terrible normals, now they are ok, so this isn't needed anymore)
 +
  -v: verbose
  
 +
note: source code in /storage/big1/lml/Fischer/src directory.
 
   
 
   
−
===  connected_random_pts ===
+
</nowiki>
−
Generates random pts in a planar region, calculates how many connected components are created
+
===  BestPath ===
−
if a pt is connected to all other pts within a given radius.
+
<nowiki>
 +
 
 +
Finds the brightest path from a start pixel to a stop pixel (1-indexed).
  
 
Usage:
 
Usage:
−
connected_random_pts [options]
+
BestPath [options] image.i2i
 
Options:
 
Options:
−
-x lowx hix:         pts have xcoords within this range, default = (0.0,100.0)
+
-t #: threshold. Only pixels with values > # are allowed in the path, default = 0.000000
−
-y lowy hiy:         pts have ycoords within this range, default = (0.0,100.0)
+
-s x y z: start coordinate (1-indexed). default = (1,1,1)
−
-r #: radius (in pixels), default = 5.0
+
-S x y z: stop coordinate (1-indexed). default = (1,1,1)
−
-s #: long int starting seed for random number generator. default = process id.
+
-v: verbose
−
-n #: number of pts to create, default = 1000
 
−
  -h: help
 
−
  -v: verbose
 
−
 
 
−
See also:
 
−
krypton/Huang/README, components.pl, Kth_pt.m, num_observed.m, mean_minimum_distance.m,objs2mst.pl
 
  
 
Source code in ~krypton/lml/facil   
 
Source code in ~krypton/lml/facil   
 
   
 
   
−
== Image Manipulation ==
+
</nowiki>
−
===  align_quadrants.pl ===
+
===  BestPath3D ===
−
This program displays the four quadrant images of a well. The user then saves 3 pairs of points
+
<nowiki>
−
(s when middle mouse is pressed). Each pair of points should be points that match in two
+
 
−
neighboring quadrants. When done, each quadrant must have one (or two) points specified.
+
Finds the brightest path from a start pixel to a stop pixel (1-indexed).
−
Points must be in pair order (see recalc_quadrant2.pl).  When done, the use must write them out
+
Converts voxels into nodes in a graph and finds the lowest cost path from the start node to the stop node.
−
via z (when middle mouse is also pressed, no confimation is given in play).   
+
The cost of each node is (maxval-imval), so bright nodes have less cost. Output in rpts format to stdout.
−
The quadrant images are then aligned based on these points and newimage.i2i is created.
+
BestPath3D always turns all (above threshold) voxels in the image into graph nodes. BestPath3D_ellipse only
 +
looks at voxels within an ellipse with a major axis connecting the start and endpts.
 +
Usage:
 +
BestPath3D [options] image.i2i > image.rpts
 +
BestPath3D_ellipse [options] image.i2i > image.rpts
 +
Options:
 +
-t #: threshold. Only pixels with values > # are allowed in the path, default = 0.000000
 +
-s x y z: start coordinate (1-indexed).  
 +
-S x y z: stop coordinate (1-indexed).
 +
Up to 1000  pairs of start/stop pixels can be specified. At least one pair must be specified.
 +
-r scale black  rescale image values so new = (old-black)/scale prior to being used
 +
-n name name of object that is placed into the rpts file.
 +
-f exfac perpfac: Only used if you are running BestPath3D_ellipse
 +
        sets principal axis a to have the specified endpts (from -s and -S), then expands it by factor exfac,
 +
                        and sets the length of the perpendicular b and c axes to be perpfac times the original axis a length
 +
defaults: exfac = 1.000000, perpfac = 1.000000
 +
-v: verbose
 +
-d: debug
 +
-D debug.i2i: debug. also write out image with 50 added to all voxels within the ellipsoid.
 +
 
 +
Examples:
 +
make_spiralim -d 200 200 30 -r 70 -p spiral_small.i2i # create a test image
 +
                -s 171 101 1  -S 170 99 30            # written out by make_spiralim
 +
BestPath3D_ellipse -v -t -1 -s 171 101 1 -S 170 99 30 -f 3 13 spiral_small.i2i > spiral.rpts
 +
note: spiral_small.i2i is a tough case, since the spiral goes very far from the start/stop points
 +
This requires a -f 3 to expand out the endpts and then a 13 to expand out perpendicularly.
 +
If you just expand out perpendicularly (e.g. -f 1 13) the ellipsoid doesn't grow faster enough
 +
near the endpts so some of the spiral data is not included in the ellipsoid. See -D option.
  
−
It will also create a file with the output from play, for future reference.
+
See also: make_spiralim, BestPath, BestPath2, make_ellipsoid, DAVE -> Stuff -> Best Path ...
−
It will be named, eg, G02_combined_play.pts. It can be passed manually to recalc_quadrant2.pl to redo
 
−
the alignment if necessary.
 
  
−
usage:
+
Source code in ~krypton/lml/facil 
−
  align_quadrants.pl [options] well newimage.i2i
+
 +
</nowiki>
 +
===  BestPath3D_ellipse  ===
 +
<nowiki>
  
 +
Finds the brightest path from a start pixel to a stop pixel (1-indexed).
 +
Converts voxels into nodes in a graph and finds the lowest cost path from the start node to the stop node.
 +
The cost of each node is (maxval-imval), so bright nodes have less cost. Output in rpts format to stdout.
 +
BestPath3D always turns all (above threshold) voxels in the image into graph nodes. BestPath3D_ellipse only
 +
looks at voxels within an ellipse with a major axis connecting the start and endpts.
 +
Usage:
 +
BestPath3D [options] image.i2i > image.rpts
 +
BestPath3D_ellipse [options] image.i2i > image.rpts
 +
Options:
 +
-t #: threshold. Only pixels with values > # are allowed in the path, default = 0.000000
 +
-s x y z: start coordinate (1-indexed).
 +
-S x y z: stop coordinate (1-indexed).
 +
Up to 1000  pairs of start/stop pixels can be specified. At least one pair must be specified.
 +
-r scale black  rescale image values so new = (old-black)/scale prior to being used
 +
-n name name of object that is placed into the rpts file.
 +
-f exfac perpfac:  Only used if you are running BestPath3D_ellipse
 +
        sets principal axis a to have the specified endpts (from -s and -S), then expands it by factor exfac,
 +
                        and sets the length of the perpendicular b and c axes to be perpfac times the original axis a length
 +
defaults: exfac = 1.000000, perpfac = 1.000000
 +
-v: verbose
 +
-d: debug
 +
-D debug.i2i: debug. also write out image with 50 added to all voxels within the ellipsoid.
  
−
examples:
+
Examples:
−
  align_quadrants.pl [options] G02 G02_combined.i2i
+
make_spiralim -d 200 200 30 -r 70 -p spiral_small.i2i  # create a test image
 +
                -s 171 101 1  -S 170 99 30            # written out by make_spiralim
 +
BestPath3D_ellipse -v -t -1 -s 171 101 1 -S 170 99 30 -f 3 13 spiral_small.i2i > spiral.rpts
 +
note: spiral_small.i2i is a tough case, since the spiral goes very far from the start/stop points
 +
This requires a -f 3 to expand out the endpts and then a 13 to expand out perpendicularly.
 +
If you just expand out perpendicularly (e.g. -f 1 13) the ellipsoid doesn't grow faster enough
 +
near the endpts so some of the spiral data is not included in the ellipsoid. See -D option.
  
−
options:
+
See also: make_spiralim, BestPath, BestPath2, make_ellipsoid, DAVE -> Stuff -> Best Path ...
−
-no_pts_output don't produce the pts file
 
−
-noblend by default the quadrant images are blended together where
 
−
they overlap. use this option to turn that off.
 
−
-print_overlap also print the amount of overlap to stderr
 
−
-debug
 
−
-debug_all
 
  
 +
Source code in ~krypton/lml/facil 
 +
 +
</nowiki>
 +
===  boundaryL  ===
 +
<nowiki>
 +
Usage:
 +
  boundary [options] original.i2i
  
−
see also:
+
Description:
−
combine_images
+
  Boundary takes an image, applys a threshold to the image,
−
recalc_quadrant2.pl
+
  rolls a ball outside then inside the object. This defines
−
/home/lml/krypton/Corvera/Raz/README 5/16/14
+
  what pixels are considered inside, outside, and boundary.
−
http://wiki.imagej.net/Grid/Collection_Stitching_Plugin
 
  
 +
  Expects:  Input image and optionally some given options
 +
  Modifies: Nothing
 +
  Returns:  Output image with the name of image with _B added.
 +
            (if input.i2i was original image input_B.i2i
 +
            is output image)
  
−
   
+
Options:
−
=== combine_images.pl ===
+
-I        : Do not roll ball inside of the cell.
 +
              (rolling the ball inside the cell results
 +
              in a smoother surface)
 +
  -r #      : Set radius to # for sphere exterior to cell.
 +
              (default = 3)
 +
-R #      : Set radius to # for sphere interior to cell.
 +
              (default = 3)
 +
-t #      : Set threshold for all z slices to > #.
 +
              (default is 10)
 +
-T filename: Use file filename for thresholds for each z slice.
 +
              (one threshold for each zslice - one threshold on a line)
 +
-X        : Restrict search in X direction.
 +
              (can be used to cap a cylinder whose long axis
 +
              is along the x axis)
 +
-Y        : Restrict search in Y direction.
 +
              (can be used to cap a cylinder whose long axis
 +
              is along the y axis)
 +
-i string  : Add string to history of output image.
 +
  -h        : Print this message.
 +
  -H        : Print this message and then some.
  
−
This program runs combine_images on all the images in the current directory.  All images which have names
+
Caveats:
−
like L##_morestuff.i2i (where L is any letter and # is any digit) will be combined;
+
  Requires lots of memory and time to run (megabytes+minutes).
−
all the A02 will be combined (to produce A02_comb.i2i), all the A03, etc.
+
 
−
The resulting image for the combination of the A02 images will be A02_tag.i2i (see -newtag option, e.g. A02_comb.i2i).
+
Notes:
−
Any images already existing in the current directory with a name like L##_stuff_oldtag.i2i will be ignored.
+
  Entire image name (with extension) is required.
 +
  Pixel values: boundary = 2, outside = 4, inside = 6.
 +
  Output image is padded on all sides by 2*radius+1 of
 +
  sphere rolled on the outside of the object.
  
−
usage:
+
Keywords:
−
combine_images.pl [options]
+
  surface location, boundary, boundaries, border
  
−
options:
+
Additional programs:
−
-newtag=something The output filename for L##*.i2i will be L##_newtag.i2i.  Default = comb
+
  histogram, shrink_wrap, createBoundary, analysis
−
-oldtag=something Any files of the form L##*_oldtag.i2i will be ignored. Default = comb
 
−
-pixsize=# pass this as -p # to combine_images program (pixel size).
 
−
Specify -pixsize=2.41 (um/pixel) for the 2.5X objective.
 
−
-reduceima=#:#:tag After creating output image, apply reduceima to it 
 
−
with -bin # # and the specified output tag on the name.
 
−
So, e.g., A02_comb.i2i will become A02_tag.i2i
 
−
-flipx flip images in X prior to combining them
 
−
-flipy flip images in Y prior to combining them (can be used at the same time as -flipx)
 
−
-x=#:#:#:# manually specified picture offsets. see combine images
 
−
-y=#:#:#:# manually specified picture offsets. see combine images
 
−
-o=# number of pixels the images overlap (ignores info in image headers). see combine_images
 
−
-f do MOT pad images at border (as of 2/3/14). see combine_images.
 
−
-remove Remove the full-size combined image (e.g., A02_comb.i2i) when done (use with -reduceima).
 
−
-dummy Just echo what would be done, but don't do anything
 
−
-verbose Echo as things are done.
 
−
-debug
 
  
−
note:
+
Last Changed: Wed Feb 10, 1993
−
combine_images (in /storage/big1/lml/Linux/bin) and reduceima (in /storage/big1/kef/Linuxbin) must be in your
 
−
path for this perl script to work.  You can check this by just typing: which combine_images  and  which reduceima;
 
−
if they are not found you need to add their directory to your PATH environment variable (echo $PATH will show you
 
−
what it is currently). How this is done varies depending upon which shell you are running. Under tcsh it would
 
−
be  setenv PATH $PATH:/storage/big1/kef/Linuxbin:/storage/big1/lml/Linux/bin .
 
  
−
example:
 
−
      cd /storage/big3/kdb/Zeiss/20130326/GEDMAT_002_PRO_SQ2_DAY14
 
−
# Combine all images in each well. The combined image will be L##_combined (-newtag=combined).
 
−
# Then reduce the combined size back down to the size of one original image;
 
−
# call the new image L##_combined_reduc.i2i (-reduceima=2:2:reduc). 
 
−
# Finally, remove the big (unreduced) combined image (-remove). 
 
−
# First do a dummy run (-dummy) to see if it is going to do what we want:
 
−
      combine_images.pl -newtag=combined -reduceima=2:2:reduc -pixsize=2.41 -remove -dummy   
 
−
# ok, now the real thing (just leave off -dummy):
 
−
      combine_images.pl -newtag=combined -reduceima=2:2:reduc -pixsize=2.41 -remove
 
  
−
see also:
+
</nowiki>
−
combine_images
+
===  calc_area  ===
−
/storage/big1/raz/GEDMAT
+
<nowiki>
−
mask_gedmat_images.pl
 
  
−
+
This program takes in a surface image and a data image and calculates:
−
===  overlap_images.pl  ===
+
1). the area of the surface (number of nonzero voxels)
 +
2). the number of data voxels (nonzero voxels in image2)
 +
3). the number of surface voxels (area) within a specified distance of a data voxel.
 +
This information is printed to stderr.
  
−
  Takes output from parsepos.pl  and calculates how the images overlap so they can then be combined easily (-create option).
+
Typically one might apply a shrink_wrap to, say, a Vinculin image of a dual
 +
labelled image pair. Then make_surface would convert this to a surface.  
 +
After this, map_to_surface would be applied to the 2 dual labelled images.
 +
Then overlap would be used to calculate the overlap of the two data sets
 +
(given some overlap range, which defaults to 1).
 +
Then this program (calc_area) is used to get the area of the surface and the
 +
amount of it which is examined using the range specified to the overlap program.
 +
This is important since to calculate the probability that the overlap was
 +
due to chance you want to know what percentage of the area was included in
 +
the overlap calculation.
  
−
overlap_images.pl [options]  
+
Usage: calc_area [options] surface.i2i data.i2i
 +
options:
 +
  -r range: range in voxels (default = 1).
 +
  -i image.i2i: also produce an output image
 +
  this image will have bit 2 set for original data voxels
 +
  and bit 1 set for surface voxels near (or on) data voxels
 +
  (so it would be a voxel with a value of 1 or 3).
  
−
options:
+
note: entire image name (with extension) is NOT required.
−
-xdim=# xdim of image in pixels, default = 1392
+
note: a - in place of an image name means stdin or stdout.
−
-ydim=# ydim of image in pixels, default = 1040
+
note: a compressed image will be automatically be uncompressed.
−
-zdim=# zdim of image in pixels, default = 25
 
−
-pixsize=# size of pixels in um, default = 0.645
 
−
-create=file.i2i actually combine the images into one big image  
 
−
-flipx flip each image around its x-axis prior to inserting it into the new image (if -create is specified)
 
−
-flipy flip each image around its y-axis prior to inserting it into the new image (if -create is specified)
 
−
-verbose
 
−
-dummy just echo what would be done, but don't do anything (nothing normally done anyway unless -create specified).
 
  
−
see also:
+
Source code in /storage/big1/lml/was_invitro/Moore/pkc
−
parsepos.pl
+
−
insert_image called by this program if -create is specified.
+
</nowiki>
−
reduceima if -create is used, this may be needed to make the image small enough to see via play.
+
===  closest_object  ===
−
+
<nowiki>
−
example:
 
−
        parsepos.pl -pre=/storage/big3/kdb/Zeiss/20121004/LectinExp3/ -post=_1_1_20121004_LectinExp3.i2i \
 
−
                /storage/big3/kdb/Zeiss/20121004/LectinExp3/LextinExp3.pos | overlap_images.pl -create=newimage.i2i
 
−
reduceima -A -bin 4 4 newimage.i2i newimage_reduced.i2i
 
  
−
  note: you may need /storage/big1/lml/Linux/bin and /storage/big1/kef/Linux/bin in your path
+
Calculates the closest distance from an object in image1 to an object in image2. Also calculates closest distance
 +
between objects within each image separately.  Prints out info about matches.  If no other object is within the
 +
maximum distance (see -m option), the object is printed matching to object -1 at a huge distance.
 +
Histogram of these distances is printed to stdout.
 +
The input images should have each pixel in an object having the object id as its value. These images can be
 +
produced with "countobjs -Q" or "objs_via_ascent" 
  
−
  source code in /storage/big1/lml/Corvera/Olga/data/ptrs
+
Usage:
−
see also: insert_image
+
closest_object [options] image1.i2i image2.i2i > im1_im2.hist
−
  http://bigwww.epfl.ch/algorithms.html -> MosaicJ (which may already exist in Fiji)
+
Options:
 +
-t # #: image1 and image2 thresholds, pixels > # are above threshold, default = 0.
 +
-w # # #: x,y, zweights for pixel distances (ie, pixel size), default = 1 for each.
 +
-m # :         maxdistance (in units used for -w) to look for the closest pixel, default = 10.000000
 +
-r #:         resolution for output histogram, ie, binsize, default = 0.100000
 +
-e lowx lowy lowz hix hiy hiz: only look at voxels within this extent (subvolume), 1-indexed, inclusive.
 +
-s minsize maxsize:    only look at objects in image1.i2i within the size range (inclusive, in voxels)
 +
-S minsize maxsize:    only look at objects in image2.i2i within the size range (inclusive, in voxels)
 +
-i miniod maxiod:    only look at objects in image1.i2i within the iod range (inclusive)
 +
-I miniod maxiod:    only look at objects in image2.i2i within the iod range (inclusive)
 +
-d image1d.i2i: the original intensity image for image1 (since image1 has pixel values as id numbers)
 +
-D image2d.i2i: the original intensity image for image2 (since image2 has pixel values as id numbers)
 +
if you want the iod or pixel intensity stats to be meaningful, you need to provide
 +
the original data by using the -d and -D options.
 +
  -v: verbose
 +
  -V: very verbose
 +
Examples:
 +
    Find the distance from the closest Ryr voxel in each object to a BK object. Record the iod
 +
    of the BK object in the histogram:
 +
countobjs -t 5 10 -Q ryr_id.i2i ryr.countobjs ryr.i2i      : this finds Ryr connected objects and labels them with id
 +
countobjs -t 10 10 -O bk_iodR.i2i bk.countobjs bk.i2i     : this calculates the iod of BK objects
 +
float2intL -M 30000 bk_iodR.i2i bk_iod.i2i     : rescale BK iod to integer format
 +
closest_object -w 1 1 3 -o 5 100  ryr_id.i2i bk_iod.i2i > ryr_bk.info
 +
cull -s -c -1 -4 < ryr_bk.info |xmgrace -source stdin
 +
        note: you may want to take a 10 pt running avg inside xmgrace to compensate for
 +
issues related to discretization of distances
 +
See also:
 +
closest_voxel objs_via_ascent /home/lml/krypton/Zhuge/BKandRyr/graph.pl,match.pl
 +
Bugs:
  
 +
Source code in ~krypton/lml/facil 
 
   
 
   
−
===  parsepos.pl ===
+
</nowiki>
−
  Parse a position file produced by Zeiss system to pull out x y coords for each of the images taken in well C09.
+
===  closest_voxel ===
−
Output triples (well_id x y) written to stdout.
+
  <nowiki>
−
Position may be in microns, x increasing to the right, y increasing down.
+
 
 +
Calculates the closest distance from an above threshold voxel in image1 to an above threshold voxel in image2.
 +
Histogram of these distances is printed to stdout.
  
−
  parsepos.pl filename.pos
+
Usage:
 +
closest_voxel [options] image1.i2i image2.i2i > im1_im2.hist
 +
Options:
 +
-t # #: image1 and image2 thresholds, pixels > # are above threshold, default = 0.
 +
-w # # #: x,y, zweights for pixel distances (ie, pixel size), default = 1 for each.
 +
-m # :         maxdistance (in units used for -w) to look for the closest pixel, default = 10.000000
 +
-r #:         resolution for output histogram, ie, binsize, default = 0.100000
 +
-s lowx lowy lowz hix hiy hiz: only look at voxels within this subvolume, 1-indexed, inclusive.
 +
-Q: only the voxel in each image1 object closest to an image2 voxel should count.
 +
Use this if image1.i2i was created via the -Q option from countobjs.
 +
-o minsize maxsize:    only look at objects in image1.i2i within the size range (inclusive, in voxels)
 +
must use -Q option along with this.
 +
-p: also print the detailed info of which voxel matched to which other voxel
 +
  -v: verbose
 +
  -V: very verbose
 +
Examples:
 +
    Find the distance from the closest Ryr voxel in each object to a BK object. Record the iod
 +
    of the BK object in the histogram:
 +
countobjs -t 5 10 -Q ryr_id.i2i ryr.countobjs ryr.i2i      : this finds Ryr connected objects and labels them with id
 +
countobjs -t 10 10 -O bk_iodR.i2i bk.countobjs bk.i2i     : this calculates the iod of BK objects
 +
float2intL -M 30000 bk_iodR.i2i bk_iod.i2i     : rescale BK iod to integer format
 +
closest_voxel -w 1 1 3 -o 5 100 -Q ryr_id.i2i bk_iod.i2i > ryr_bk.info
 +
cull -s -c -1 -4 < ryr_bk.info |xmgrace -source stdin
 +
        note: you may want to take a 10 pt running avg inside xmgrace to compensate for
 +
issues related to discretization of distances
 +
See also:
 +
graph.pl  
 +
Bugs:
  
−
  options:
+
Source code in ~krypton/lml/facil 
−
-well=string well to analyze. default = C09
+
   
−
-pre=prestring add this string in front of the well_id
+
</nowiki>
−
-post=postring add this string after the well_id
+
=== coloc  ===
−
these two options are needed if you want to pass this output to
+
<nowiki>
−
overlap_image.pl -create=newimage.i2i, since that will need to be able
 
−
to find the image associated with each well_id.
 
−
-verbose additional output to stderr
 
−
-vverbose additional output to stderr
 
  
−
  example:
+
  This program reads in 2 or 3 images, and writes out an image in which
−
parsepos.pl -pre=/storage/big3/kdb/Zeiss/20121004/LectinExp3/ -post=_1_1_20121004_LectinExp3.i2i  \
+
colocalized voxels (voxels > threshold in all images) are kept, others
−
/storage/big3/kdb/Zeiss/20121004/LectinExp3/LextinExp3.pos | overlap_images.pl -create=newimage.i2i
+
set to 0.  By default colocalized voxels are set to image1 values.
−
+
Usage: coloc [options] image1 image2 [image3] outimage
 +
options:
 +
-a #:        threshold for image 1 (default = 0)
 +
-b #:        threshold for image 2 (default = 0)
 +
-c #:        threshold for image 3 (default = 0)
 +
-va:          make outimage pixel value be value of image 1
 +
-vb:          make outimage pixel value be value of image 2
 +
-vc:          make outimage pixel value be value of image 3
 +
-vu #:        make outimage pixel value be user specified, #
 +
note: source code in /storage/big1/lml/Laxman/src/coloc.c
  
 +
</nowiki>
 +
===  coloc3ims  ===
 +
<nowiki>
  
−
see also:
 
−
overlap_images.pl
 
−
insert_image
 
  
−
  note: you may need /storage/big1/lml/Linux/bin and /storage/big1/kef/Linux/bin in your path.
+
This programs reads in three images and produces a new version of image3.
 +
Each voxel which is colocalized (has image1 > thresh1
 +
and image2 > thresh2) has the value in image3 set to 32000 (-n 32000). Otherwise the value in image3 is not
 +
changed. Once these modifications are made, image1.i2i image2.i2i and newimage3.i2i can be read
 +
into DAVE (in that order) and voxels which are colocalized in the first two channels
 +
will be guaranteed to be colocalized in the third channel.
 +
 
 +
Usage: coloc3ims [options] image1.i2i image2.i2i image3.i2i newimage3.i2i
 +
options:
 +
  -t thresh1 thresh2:  thresholds for image1 and image2, defaults = 0 and 0
 +
  -n newval: instead of setting newimage3 to 0, set it to newval
 +
  -m: instead of setting newimage3 to 0, set it to the max value in image3.i2i
 +
 
 +
note: entire image name (with extension) is NOT required.
 +
note: a - in place of an image name means stdin or stdout.
 +
note: a compressed image will be automatically be uncompressed.
 +
 +
</nowiki>
 +
===  coloc_tseries2  ===
 +
<nowiki>
 +
 
 +
Similar to coloc_tseries, but compares a 2D time series to itself.
 +
Prints xmgr formatted graphs to stdout.
 +
 
 +
Usage: coloc_tseries2  image1 outfile
 +
options:
 +
-t thresh:    only count voxels > thresh. default = 0
 +
-z start stop: only look at these z planes (inclusive, 1-indexed). default = all z planes.
 +
-n: don't include a legend in the stdout.
 +
        -v:          verbose
 +
 
 +
source code in:  /home/lml/krypton/facil
 +
 
 +
see also: /home/lml/krypton/Corvera/paper2010/Dynamics_in_Dave/README
 +
 +
</nowiki>
 +
===  com_2d_objs  ===
 +
<nowiki>
 +
 
 +
This program takes an image and finds the intensity weighted center of mass of each 2d object.
 +
Object 1 is all pixels with value = 1, object 2 is pixels with value 2, etc. Max objects = 10
 +
Statistics are written to stdout
 +
 
 +
Usage: com_2d_objs [options] image.i2i
 +
options:
 +
  -z start stop: zplanes to analyze. 0-indexed inclusive. default is all z planes
 +
  -r #: put data out rpts format (integer coords, only center of mass info). Only use if only one object.
 +
# is the pixel value (!= 0), usually -1 through -7.
 +
  -o: coords should be one-indexed (ie, min x = 1). default = 0. -r sets this automatically.
 +
  -v: verbose
 +
 
 +
Source in /home/lml/krypton/facil
 +
See Also: /home/lml/krypton/Lawson/Jacques/README
  
 +
Examples:
 +
  planimeter3.20081007 -I 110215ligand_0_491.i2i -R 110215ligand_0_491_cell1.rpts
 +
  interp -S -p -1 -o cell1 110215ligand_0_491_cell1.rpts 110215ligand_0_491_cell1_interpS.rpts
 +
  create_dcimL -d 1 -s 502 501 1440 one.i2i
 +
  addlines one.i2i 110215ligand_0_491_cell1_interpS.rpts - | maskedsmul -O 0 one.i2i - cell1_binary.i2i
 +
    # could do the same for a second object (etc), with create_dcimL -d 2, then use mathi2i
 +
    # to add the images before analysis
 +
  com_2d_objs cell1_binary.i2i > cell1_binary.stats
 
   
 
   
−
===  prep_and_restore.pl ===
+
</nowiki>
 +
===  exo4 ===
 +
<nowiki>
 +
Phase 1: Associates each voxel in inimage.i2i which is > threshold with a maxima in the image.  It finds
 +
        the maxima by doing a steepest ascent walk in the image.  Writes summary info about regions found to
 +
        stdout in the same format as countobjs.  Only the location of the maxima found in phase 1 are passed to phase 2.
 +
        All vesicles must start out as maxima. (see also -readmax option)
 +
Phase 2: In each rolling window of time points (see -w), examines the intensity over time of each maxima
 +
        (found in previous step) to see if it
 +
        meets certain criteria (i.e., looks like a vesicle fusing with the plasma membrane - undergoing
 +
        exocytosis).  If it matches this criteria it prints out the "path" of the maxima and its
 +
        pixel value and iod value.  These paths can be converted to rpts format for
 +
        addlines or xmgr format
 +
        using found4.pl. 
 +
The fusion model: think of intensity as shown below. once the vesicle primes its intensity jumps. when it fuses
 +
it drops back down.
  
−
Restores all the i2i files in the current directory. Skips any files with   _.i2i or _r.i2i in their name.
+
peak intensity profile:
−
Also ignores any files which do not have dat or ctx in their name.
+
   pintens                  ----------------
 +
                          |                |
 +
                        |                  |
 +
                        |                    |
 +
  tintens -------------                      ------------------
  
−
options:
+
          0  t1       t2                  t3
−
-dummy just print what would be done
+
          0  t1      |prise|              |ffall|
−
-bkgnd=# pass this through to find_bkgnd as the -a option. eg, -bkgnd=-.10 will lower the bkgnd value
 
−
which is subtracted off by 10 percent.
 
−
        -smooth=#       smoothness factor (smaller is less smoothing). default = .00002
 
−
        -iterations=#  maximum number of iterations, default = 999
 
−
        -scale=#:#      GFP and RFP output scaling for restoration (respectively). default = 1.0 and 1.0.
 
−
-verbose echo what is being done
 
−
        -maxfiles=# only analyze this many files. primarily for debugging
 
−
-twod during background subtraction, vary the background value for each 2D slice, instead of
 
−
just using the same value for all z slices.
 
−
-files=:file.i2i:...:filen.i2i   just restore these files. and do it even if _.i2i or _r.i2i exists.
 
−
-dont_restore  do the data prep (filename_.i2i and filename__.i2i produced), but don't do the restoration (but not filename__r.i2i).
 
−
-beowulf queue the restorations to run on our beowulf (dispatched via qsub, using pbs (torque), from a daemon running on m13)
 
−
rather than immediately on the localhost.
 
−
see also: pbsnodes (on m13, only servers running fc16 or fc17 are on the allowed list right now)
 
−
          qstat -f (on m13) will show lots of info on each job.
 
−
If this is not specified, each restoration is run sequentially, with this script blocking while waiting for each one.
 
−
-gpu only use this if you are logged into zirconium. It will run the restoration on zirconium using the GPU version
 
−
of the restoration.
 
−
-outdir=dir    rather than place the results in the current directory, place it in the fully qualified specified directory.
 
−
leave off the last /
 
−
      -GFPpsf=file    fully qualified path to GPFpsf (used when file name is *_#_#_1*.i2i, eg,  file_0_1_1.i2i)
 
−
default = /storage/big1/lml/Corvera/Olga/data/GFPpsf_segxyz_.i2i
 
−
note: psf should have background values of ~0, otherwise 0-padding will produce blocking artifacts.
 
−
      -RFPpsf=file    fully qualified path to RPFpsf (used when file name is *_#_#_2*.i2i, eg,  file_0_1_2.i2i)
 
−
default = /storage/big1/lml/Corvera/Olga/data/RFPpsf_segxyz_.i2i
 
−
note: psf should have background values of ~0, otherwise 0-padding will produce blocking artifacts.
 
−
-silent print less info
 
  
 +
t1: time of tethering (should stop moving)
 +
t2: time of priming (peak intensity increases from tintens to pintens)
 +
t3: time of fusing (peak intensity drops back down, but iod should stay high and possibly even increase a bit)
 +
prise is the time it takes to rise froom tintens to pintens during priming (docking)
 +
ffall is the time it takes to fall from pintens to tintens during fusion.
 +
 +
exo4 is similar to exo3 but has slightly different options and checks all possible fusion times
 +
simultaneously. exo3 only picked the latest (in time) possible fusion event based only upon intensity
 +
changes during the falltime. exo4 considers what is happening elsewhen in the window before deciding
 +
on a fusion time. It picks the fusion candidate (iod rise and intensity fall) which is closest in time to
 +
the maxima disappearing (-check_max) and which has a stationary period (-primedtime) within the
 +
specified time range. Hopefully this makes it less likely to make mistakes.
  
−
  examples:
+
Should work in 2d or 3d.  Only reads in a window of time points at once
−
cd /storage/big3/kdb/Zeiss/20121004/LectinExp3
+
so should be able to handle large images.
−
        /home/lml/krypton/Corvera/Olga/prep_and_restore.pl -files=G11_0_1_1_20121004_LectinExp3.i2i:G03_0_1_2_20121004_LectinExp3.i2i
 
−
      -beowulf -outdir=/storage/big1/lml/Corvera/Olga/data
 
  
−
  see also:
+
Usage:
−
epr_beowulf   a GUI front end for submitting individual restoration jobs to the beowulf queue
+
   exo4 [options] inimage.i2i  > inimage.count
−
/storage/big1/epr/bin/cuda/epr_gpu -cuda 1  if logged into zirconium, will run a single job on the graphics card (like -gpu option).
 
−
prepdata
 
−
preppsf
 
  
−
  note: /storage/big1/lml/Linux/bin /storage/big1/lml/Linux/bin64 and /storage/big1/kef/Linux/bin probably need to be in your
+
Options:
−
path to successfully run this script.
 
  
−
   
+
options related to input:
−
=== prepfiles.pl  ===
+
  -T startt stopt : only examine these time points (0-indexed, inclusive)
−
prepfiles.pl [options]  
+
  -p numskip numuse : patterned read. skip numskip z planes, then analyze numuse zplanes, then repeat.
 +
  This is useful if the acquisition protocol was, e.g., 1 fps for 1 min, 20 fps for 3 seconds
 +
  repeated over and over, and you only want to look at the 20 fps data.
 +
  numuse must be >= window size (see -w option). Pattern starts at t=0 even if -T is
 +
  specified (but startt time points are still skipped).
 +
  -w #         : number of time pts in a window, default = 100
 +
                        a negative number means read in entire dataset at once (one big window)
 +
  -jump # : jump # time pts before examining next window of time pts. This doesn't skip data, just
 +
  analyzes a "window" less often (so more has changed). Default = 0
 +
                        which means the very next time pt is added to window (and oldest dropped) and then
 +
                        the window of data is re-evaluated. May not work with -p option.
 +
  -r rptsfile : read in rptsfile.  should have one closed object in it. Uses this as a mask for all the planes.
 +
  -d tdim : the time dimension of the image (over-rides anything stored in the header)
 +
  -b #: : blur (smooth) image before looking for objects (maxima), using a window of the specified
 +
: halfwidth (in pixels) in x,y, and z (z width will be set to 0 if 2*width > zdim).
  
−
Takes a dual-wavelength image and splits it into two separate images (segim_huge), applies scratchng, and prepdata.
+
Phase 1: options related object finding via ascent (see objs_via_ascent2d), info printed out.
−
Run this from a directory. It will be applied to all images in the directory. There must be
+
          If an object doesn't satisfy these criteria, its maxima is not kept, so it is not considered a maxima
−
a file called dark.i2i in the directory too. Removes all intermediate files and original 2wavelength image.
+
          for the next stage of processing which tracks maxima into paths and checks if they are fusion events.
 +
          The iod and size specified here are also used later, but the next phase just looks with the square iod region.
 +
  -t threshold : any voxel > threshold in inimage.i2i is tracked to a 3d maxima (default = 0.0).
 +
  -i miniod maxiod : erase all objects whose iod doesn't fall in the specified range (default: 1-1000000000)
 +
  -s minsize maxsize : erase all objects whose size (in voxels, inclusive) doesn't fall in the specified range
 +
  (default = 1-1000000). note: object size and iod might be large since all voxels which
 +
  connect up via steepest ascent algorithm to the maximum count in this first phase.
 +
  -strict_max : ignore "maxima" which are really on a plateau (option might cause
 +
                        saturated maxima to be missed)
 +
  -readmax file : skip phase 1, instead just reading maxima in from the specified file
  
−
options:
+
Phase 1b: noptions related to fitting Gaussians (not used for anything yet):
−
   -safe : don't delete the original dual-wavelength image
+
   -g # : fit Gaussians (+ dc term) to each object found (eventually I will use this for something).
−
   -echo : echo what is being done
+
  only includes pixels in object within # pixels from the brightest point.
 +
  -G # : same as -g, but uses gradient info of fuction during fit (i.e., a different fit routine).
 +
  The -G version seems a bit buggy.
 +
   -n : normalize the Gaussian fom (shouldn't affect individual fits, just allow comparison between fits
  
−
   
+
Phase 2: options related to "paths" over time:
−
=== recalc_quadrant2.pl  ===
+
  Maxima must exist for 0 time pts, be stationary for 0 time pts prior to disappearing
−
align_quadrants.pl calls this program. It is easier to use it than to call recalc_quadrant2.pl manually.
+
  While a maxima is present, a pixel within range only counts as part of object
 +
  if above threshold (-t option). While a maxima is present,
 +
  a region will only be considered if size (pixels > thresh and within range) is within size (-s)
 +
  and iod is within iod (-i option).
 +
  -iodsize #           check iod within +/- range pixels., default = 3
 +
  -maxspeed #   max distance a maxima can move (in pixels) between time points, default = 5
 +
  -diststill #          how far (in x or y) in pixels a maxima can move and still be considered "still",
 +
  (this is the location compared to last position of maxima) default = 0
 +
  -fall time frac   maximum # of time pts peak intensity can take to show the specified fractional decrease
 +
  when vesicle is fusing. default = 10 and 0.300000
 +
  -iodrise frac   the fractional increase in iod which must be seen
 +
  when vesicle is fusing (see -fall). default = 0
 +
  -check_max #   vesicle must also change from max to not a max within # timepts after fusing
 +
  -minpeakval #          peak intensity value must exceed this when vesicle is fusing (-fall time parameter)
 +
  -primedtime # #   minimum and max # of time pts vesicle must be primed before fusing.
  
−
This program converts from pts printed by play (so in the coordinate system gotten from combining the four quadrant images of a well (with no overlap))
+
options related to output:
−
to one used by register_images.pl (ie, each pt is in the coordinate system of its own individual image).
+
  -debugpath x y z      print out debug info about the path that passes near (within 5 pixels) here
−
It also pairs the points up and orders them the way that register_images.pl requires.
+
                        (fgrep output for the word: debugpath). can be used 10 times. 1-indexed.
−
It assumes the user picked matching pairs of pts in order when using play. Eg, first a pt was picked (s key while holding down middle mouse button),
+
  A -1 for z, means debug any path near x and y, regardless of z value (only if -L used).
−
then its matching point in a neighboring quadrant was picked. This was then repeated 3 times (so 3 pairs of points are present).
+
  -neardebug #        for -debugpath, definition of near, in pixels, default = 5
−
note: the file names for the files associated with each quadrant are not stored in the output by default, just the quadrant number,
+
  -onlydebug      : only print out info for debug paths. ignore all other paths.
−
      use -qnames to change this; they are needed if you want to subsequently have register_images.pl use chained.txt to actually combine the images.
+
  -a : print all info about each object found (like countobjs does) rather than just summary stats
 +
                      : for each time point.
 +
  -o outimage : create an ouput image of marked objects (object ids reused each time point). Max of each
 +
: object will be marked with the negative of the object id.
 +
  -fit_model : fit a square wave to intensity profile of each vesicle found to be fusing (and to debugpaths)
 +
                        not used for anything yet.
 +
  -v : verbose
 +
  -V : very verbose. may produce huge files. put this as first option to get option parsing info out.
  
 +
Examples:
 +
max2d_simple -t 50 inputimage.i2i -p max.pts
 +
exo4 -v -r mask.rpts -d 600 -b 1 -w 100 -jump 20
 +
  -strict_max -readmax max.pts  -s 10 1000 -i 10 1000000 -t 1
 +
      -duration 20 -timestill 20 -maxspeed 5 -iodsize 5 -diststill 2
 +
      -rise 25 .3 -fall 15 .3 -iodrise .1 -minpeakval 60 primedtime 30 90
 +
      -neardebug 2 -debugpath 152 161 237 -debugpath 134 124 -1 
 +
      inputimage.i2i >& inputimage.out
 +
graph_exo3.csh inputimage.out,  which does the following:
 +
    findpaths_exo3.pl -trackxmgr=tmpavg.xmgr inputimage.out
 +
    findpaths_exo3.pl -trackxmgr=tmppeak.xmgr -peak inputimage.out
 +
    findpaths_exo3.pl -trackxmgr=tmpfit.xmgr -finalfit inputimage.out
 +
    findpaths_exo3.pl -trackxmgr=tmpdist.xmgr -distance inputimage.out
 +
    xmgr -graph 0 tmppeak.xmgr tmpfit.xmgr -graph 1 tmpavg.xmgr -graph 2 tmpdist.xmgr &
 +
in xmgr: Edit->Arrange Graphs, select all the graphs, then set rows to 3,
 +
        Edit-Preferences set Crosshair cursor, linked scrolling.
 +
findpaths_exo3.pl -trackrpts=tmp.rpts -printfusion=4 inputimage.out
 +
addlines inputimage.i2i tmp.rpts - | play -P -1 R -P -2 G -
 +
dave -n 0 -u -K -I inputimage.i2i tmp.rpts
  
−
recalc_quadrant2.pl [options] < pts.txt > chained.txt
+
source code in /home/lml/krypton/facil
 +
see also:
 +
  objs_via_ascent,  ~lml/krypton/Huang/TIRF4/found4.pl, run7
 +
#/home/lml/krypton/bin/exo4
  
−
options:
 
−
-res=# resolution the display was downsampled by prior to getting coords (ie, what was used in chres, see below).
 
−
if chres was .5, then -res=2, ie, 1/chres value. default = 2.
 
−
-xdim=# default = 1392
 
−
-ydim=# default = 1040
 
−
-qnames=name0:name1:name2:name3    file names associated with the images used to produce each of the 4 quadrants.
 
−
  they must be in this order. If not specified, then the output will not be
 
−
  usable by register_images.pl (unless -root is implemented there).
 
−
-outimage=image.i2i also produce an output combined image (ie, call register_images.pl, see below).
 
−
-qnames required for this option.
 
−
-blend if -outimage is specified, then blend the overlapping images to produce the output image.
 
−
-verbose print overlap info to stderr. maybe to use in -o option to combine_images.
 
−
-debug stuff to stderr
 
  
−
more info. the quadrants in the combined image (see combine_images below) are numbered as follows:
+
</nowiki>
−
quadrant 1 |  quadrant 3
+
===  exo6 ===
−
quadrant 0  |  quadrant 2
+
<nowiki>
−
    where x increases to the right, y increases up
+
This programs find vesicle fusion events in a 2D time series.
  
−
example:
+
Each maxima (-readmax) gets a size and iod associated with it.  Only pixels > threshold (-t)
 +
within the iod region (-iodsize) count toward the iod and size.  Once the maximum disappears
 +
(-nomax_length) the region centered around the previous location of the maximum is used.
 +
Maxima are tracked from one time pt to the next by finding the closest maxima (within -maxspeed)
 +
to its previous location.  Tracks continue after the maximum disappears for -nomax_length time pts.
  
−
# combine the 4 well images without overlap to see what pts might match between quadrants:
+
At the end of this time, the path is then analyzed to see if it meets criteria for a fusing vesicle.
−
    combine_images -o 0 F06_0_2_0_20140505_GEDMAT_P025_WES_OM_Day14.i2i F06_1_2_0_20140505_GEDMAT_P025_WES_OM_Day14.i2i 
+
All time pts during which the maxima exists must have an iod within the correct range (-i) and
−
          F06_2_2_0_20140505_GEDMAT_P025_WES_OM_Day14.i2i F06_3_2_0_20140505_GEDMAT_P025_WES_OM_Day14.i2i tmp.i2i
+
and correct size (-s). The fusion time is identified by the intensity of the maxima falling
 +
fast enough and far enough (-fall); while within the same interval the iod rises enough (-iodrise).
 +
Fusion time is then further refined to be the time of largest iod increase (between adjacent time pts)
 +
within this interval.
  
−
# look at the resulting image and identify matching pts:
+
A fusion event may also need to be the correct time interval from when the maxima disappears (-check_max).
−
    chres -S .5 .5 1 tmp.i2i - |play -     # save (s) pairs of pts which match between the quadrants. print them using z.  create file F06_pts.txt:
+
It may also need to be significantly brighter than the background (-minpeakratio).
−
marked points
+
It may be required to have a minimum peak value during fusion (-minpeakval).
−
x= 769 y= 497 z=  1 set=  1 value=    3099
 
−
x= 799 y= 525 z=  1 set=  1 value=    3684
 
−
x= 709 y= 819 z=  1 set=  1 value=    4039
 
−
x= 682 y= 854 z=  1 set=  1 value=    3974
 
−
x= 518 y= 497 z=  1 set=  1 value=    3257
 
−
x= 543 y= 524 z=  1 set=  1 value=    3436
 
  
−
# now use this program to multiply by 2, subtract xdim (1392) and/or ydim (1040) as appropriate, pair and order pts.
+
Once the fusion time is found, the program finds the stationary (-diststill) time interval prior to fusion.
−
    recalc_quadrant2.pl -outimage=F06_combined.i2i -qnames=F06_0_2_0_20140505_GEDMAT_P025_WES_OM_Day14.i2i:F06_1_2_0_20140505_GEDMAT_P025_WES_OM_Day14.i2i:F06_2_2_0_20140505_GEDMAT_P025_WES_OM_Day14.i2i:F06_3_2_0_20140505_GEDMAT_P025_WES_OM_Day14.i2i  < F06_pts.txt > F06_chained.txt
+
This (priming) time interval must be within certain constraints (-primedtime) for the path to be considered
 +
representing a valid fusion event.
 +
 +
exo6 is similar to exo4 but reads in one time point at a time, there is no concept of a "window"
 +
It only works on a 2D time series
 +
 
 +
Usage:
 +
  exo6 [options] -readmax max.rpts inimage.i2i  > inimage.count
 +
 
 +
Options:
 +
 
 +
options related to input:
 +
  -T startt stopt : only examine these time points (0-indexed, inclusive)
 +
  -r rptsfile : read in rptsfile.  should have one closed object in it. Uses this as a mask for all the planes.
 +
  -d tdim : the time dimension of the image (over-rides anything stored in the header)
 +
  -b #: : blur (smooth) image before looking for objects (maxima), using a window of the specified
 +
: halfwidth (in pixels) in x,y, and z (z width will be set to 0 if 2*width > zdim).
 +
  -readmax file : read maxima in from the specified file (this is REQUIRED)
 +
 
 +
options related to finding fusion events:
 +
  -t threshold : any voxel > threshold in inimage.i2i is tracked to a 3d maxima (default = 0.0).
 +
  -i miniod maxiod : erase all objects whose iod doesn't fall in the specified range (default: 1-1000000000)
 +
  -s minsize maxsize : erase all objects whose size (in voxels, inclusive) doesn't fall in the specified range
 +
  (default = 1-1000000). note: object size and iod might be large since all voxels which
 +
  connect up via steepest ascent algorithm to the maximum count in this first phase.
 +
  -iodsize #           check iod within +/- range pixels., default = 3
 +
  -maxspeed #   max distance a maxima can move (in pixels) between time points, default = 5
 +
  -diststill #          how far (in x or y) in pixels a maxima can move and still be considered "still",
 +
  (this is the location compared to last position of maxima) default = 0
 +
  -fall time frac   maximum # of time pts peak intensity can take to show the specified fractional decrease
 +
  when vesicle is fusing. default = 10 and 0.300000
 +
  -startfall time   how close (in timepts) in time the peak intensity must start to fall in relation
 +
  to the fusion time (biggest jump in iod between two consecutive time pts). default = 1
 +
  -iodrise frac   the fractional increase in iod which must be seen
 +
  when vesicle is fusing (see -fall). default = 0
 +
  -check_max min# max#   vesicle must also change from max to not a max >= min# timepts and <= max# timepts after fusing
 +
  -nomax_length #        only analyze a path # time pts after max first disappears
 +
                        the default (which is also the minimum allowed value) = 1
 +
  -minpeakval #          peak intensity value must exceed this sometime while vesicle is fusing (-fall time  parameter)
 +
  default value = 0
 +
  -minpeakratio ratio# time# dist#:   the ratio of maxval/(median bkgnd value) must >= ratio# within
 +
  time# time steps of fusion.
 +
  bkgnd values are those pixels whose distance from max is >= dist# and <= iodsize
 +
  defaults: not to check this at all
 +
  -primedtime # #   minimum and max # of time pts vesicle must be primed before fusing.
 +
 
 +
options related to speed:
 +
  -nomedian   if -minpeakratio is defined, this causes the avg bkgnd value to be used instead
 +
  of the median value (calculation of median can be expensive).
 +
  -buffered_read #   read # z planes at a time to speed up disk I/O. Default is 1 z plane at a time
  
−
# same thing, but in two steps.
+
options related to output:
−
    recalc_quadrant2.pl -qnames=F06_0_2_0_20140505_GEDMAT_P025_WES_OM_Day14.i2i:F06_1_2_0_20140505_GEDMAT_P025_WES_OM_Day14.i2i:F06_2_2_0_20140505_GEDMAT_P025_WES_OM_Day14.i2i:F06_3_2_0_20140505_GEDMAT_P025_WES_OM_Day14.i2i  < F06_pts.txt > F06_chained.txt
+
  -debugpath x y z      print out debug info about the path that passes near (within 5 pixels) here
−
    register_images.pl -zdim=5 -xscale=1 -yscale=1 -post= -debug -out=F06_combined.i2i F06_chained.txt
+
                        (fgrep output for the word: debugpath). can be used 10 times. 1-indexed.
 +
  A -1 for z, means debug any path near x and y, regardless of z value (only if -L used).
 +
  -neardebug #        for -debugpath, definition of near, in pixels, default = 5
 +
  -onlydebug      : only print out info for debug paths. ignore all other paths.
 +
  -a : print all info about each object found (like countobjs does) rather than just summary stats
 +
                      : for each time point.
 +
  -o outimage : create an ouput image of marked objects (object ids reused each time point). Max of each
 +
: object will be marked with the negative of the object id.
 +
  -v : verbose
 +
  -V : very verbose. may produce huge files. put this as first option to get option parsing info out.
  
 +
Examples:
 +
max2d_simple -t 50 inputimage.i2i -p max.pts
 +
exo6 -v -r mask.rpts -d 600 
 +
  -readmax max.pts  -s 10 1000 -i 10 1000000 -t 1
 +
      -timestill 20 -maxspeed 5 -iodsize 5 -diststill 2
 +
      -fall 15 .3 -iodrise .1 -minpeakval 60 primedtime 30 90
 +
      -neardebug 2 -debugpath 152 161 237 -debugpath 134 124 -1 
 +
      inputimage.i2i >& inputimage.out
 +
graph_exo3.csh inputimage.out,  which does the following:
 +
    findpaths_exo3.pl -trackxmgr=tmpavg.xmgr inputimage.out
 +
    findpaths_exo3.pl -trackxmgr=tmppeak.xmgr -peak inputimage.out
 +
    findpaths_exo3.pl -trackxmgr=tmpfit.xmgr -finalfit inputimage.out
 +
    findpaths_exo3.pl -trackxmgr=tmpdist.xmgr -distance inputimage.out
 +
    xmgr -graph 0 tmppeak.xmgr tmpfit.xmgr -graph 1 tmpavg.xmgr -graph 2 tmpdist.xmgr &
 +
in xmgr: Edit->Arrange Graphs, select all the graphs, then set rows to 3,
 +
        Edit-Preferences set Crosshair cursor, linked scrolling.
 +
findpaths_exo5.pl -rptsfile=tmp.rpts -rptstracks -printfusion=4 inputimage.out
 +
addlines inputimage.i2i tmp.rpts - | play -P -1 R -P -2 G -
 +
dave -n 0 -u -K -I inputimage.i2i tmp.rpts
  
 +
source code in /home/lml/krypton/facil
 
see also:
 
see also:
−
/home/lml/krypton/Corvera/Raz/README 5/15/14
+
  objs_via_ascent,  track_back2, ~lml/krypton/Huang/TIRF4/findpaths_exo5.pl, run7, graph_exo3.pl
−
align_quadrants.pl
+
#/home/lml/krypton/bin/exo6
−
recalc_quadrant.pl
+
 
−
combine_images
+
 
−
combine_images.pl
+
</nowiki>
−
register_images.pl
+
=== find_2d_objs  ===
−
insert_image2
+
<nowiki>
 +
 
 +
For each z slice (time point) in the image, find 8-connected objects.  Print info to stdout.
 +
Only voxels > threshold can be part of an object (default = 0.000000). Doesn't use much memory (reads in a slice at a time).
 +
Should handle our regular (short int) image format and floating point images.
 +
 
 +
Usage:
 +
find_2d_objs [options] image1.i2i
 +
Options:
 +
-s min max: object must be within this size in pixels (inclusive), default = 1 100000
 +
-b min max: object IOD must be within this range (inclusive), default = 1.000000 1000000000.000000
 +
-t #: threshold for image1.i2i default = 0.000000
 +
-T #: adaptive threshold. # = pixel intensity/mean intensity for each time pt. must be > # to be in object.
 +
If -o specified, only pixels under the mask counted calculating mean intensity.
 +
-i thresh.i2i: use with -T option if desired. thresh.i2i is used to calculate the threshold intead of image1.i2i
 +
If -o specified, then that mask will also be applied to thresh.i2i.
 +
-n image.i2i: write out new image with all voxels not part of an object set to 0
 +
-z low high: only analyze z slices from low to high (1-indexed, inclusive).
 +
      -o mask.i2i:    apply mask image to before analyzing it. only keep pixels in image1.i2i under mask pixels > 0.
 +
mask.i2i must be in our standard image format (short int). If 3d, only first 2d image used.
 +
  -v: verbose
 +
 
 +
See also: pt_distances, max2d, max3dL, maxima, maxL, maxzL, float2intL, int2float, playexact
 +
            track, max2d_simple, countobjs
 +
 
 +
Example:
 +
        # find all objects between 5 and 30 pixels, minimum iod of 200, above a threshold of 100, write out new image:
 +
    find_2d_objs -t 100 -s 5 30 -b 200 1e9 -n newimage.i2i image.i2i > image.objs 
  
 +
Source code in ~krypton/lml/facil 
 
   
 
   
−
===  register_images.pl ===
+
</nowiki>
 +
===  find_blobs ===
 +
<nowiki>
 +
/home/lml/krypton/bin/find_blobs
 +
insufficient number of arguments
 +
This program pads an image in z, then calls "invariant" (with -F 10) to produce multiple resolutions
 +
of the original image along with its 2nd derivative in the gradient direction.
 +
The blurred originals are used to find the location of the maxima. The derivative images should
 +
have a zero crossing at the inflection points (eg, "edges"). The maxima should be in regions with
 +
negative second derivative. This program then calls flood2zero to find the spatial extent of each
 +
"blob" (bright region surrounding the maxima). Ideally this should just be a flood fill until
 +
the zero crossing is found, but noise, etc. means that a lower threshold works better.
  
−
Takes a bunch of images taken in one well on the microscope, translates them
+
USAGE: find_blobs [options] inputimage.i2i outputroot
−
based on ChainedTrans.txt and combines them into one big image.
 
  
−
ChainedTrans.txt should be a partially ordered file with entries like
+
output extensions of files produced (in same directory as inputimage.i2i):
−
X02:X03    (685,282):(736,287)  
+
      .ppts          padded points coords (location of maxima in blurred, but not deriv, image).
−
This means that location (685,282) in file X02 is the same as location (736,287) in file X03.
+
      .pts            unpadded points, with z value transformed back to unpadded coordinates
−
By partially ordered I mean that for file1:file2  file1 should already have been the
+
      .bpts          unpadded points after filtering to keep bright points (\>= min_intensity)  
−
right hand side of a similar line previously (so its position is already defined relative to
+
      .dpts          bpts with "unordered" -> "ordered" and "break" -> "#moveto", for display in DAVE
−
the file on the left hand side), unless it is the very first line in
+
      _bres#.i2i      blurred, but no derivatives, at resolution level = # (bigger # = lower res.)
−
the file. So every right hand side is relative to a left hand side which has already
+
      _dres#.i2i      blurred & derivatives taken, at resolution level = #
−
had its position defined.
+
      _dres#_obj.i2i  objects found in *_dres#.i2i by floodfilling from points in *.bpts
−
Output goes to new_trans.i2i
+
      _dres#.obj      statistics on *_dres#_obj.i2i
 +
      _flood.out      output from flood2zero
 +
      .out            output from invariant
  
−
register_images.pl [options] ChainedTrans.txt
 
−
-out=filename.i2i  name of output file produced. default = newimage.i2i
 
−
-post=string the rest of the filename after what is in ChainedTrans.txt, eg, if the file is named
 
−
X01_0_0_1.i2i, -post=_0_0_1.i2i (this is also the default for post if not specified).
 
−
-xdim=# size of original each original image (before recaling by maxtrima, see example below). default = 1392.
 
−
-ydim=# default = 1040
 
−
-zdim=# default = 21
 
−
-xscale=# binning done by matrixima (see example), coords will be scaled by this. default = 4.
 
−
-yscale=#
 
−
-blend when combining images, blend the values where they overlap (default is to just replace any previous
 
−
value with the new value).
 
−
    -dummy
 
−
-debug
 
−
-h
 
  
 +
options: 
 +
    -m #:   any maxima less than this will get filtered out, default = 1
 +
    -d #:   any derivative voxel with a value >= # will stop the flood fill
 +
                  this is in the original, floating point coord system (prior to 
 +
                  rescaling to write out deriv. image). default = -5
 +
    -p #:   pad data in z so that we don't have wrap around problems, default = 10
 +
    -z #:   number of z planes (in original image) to skip in front and back when
 +
                finding blob voxels - the derivative calculation tends to create negatives
 +
                  in the first and last few planes. default = 2 
 +
    -r #:   number of resolutions to calculate, default = 1
 +
    -e #:   maximum spatial extent of object in pixels (ie, radius), default = 5
 +
                    this is needed since the floodfill can otherwise escape and fill huge
 +
                    volumes (the first and last z slices tend to be all negative, along with
 +
                    negative values in the background regions).
 +
    -s #:    maximum number of voxels in object, default = 300
 +
    -N #:   normalize image prior to use so its max = #, default = 10000
 +
    -b sigx sigy sigz:  standard deviation,in pixels, of Gaussian used in blurring
 +
                    (passed as args for -s option of invariant), default = 2 2 1
  
−
-shift no circular shift (see tranima). old.
+
examples:
−
note: tranima will pad images with -32768 which play will show in red (I think) and DAVE will be bothered by.
 
  
−
example:
+
    find_blobs control3cluster4r_r_cell.i2i c3c4F10
−
matrixima -row 4 W X Y Z -col 3 01 02 03 -pos 0 -obj 0 -fil 1 new.i2i
+
    dave -n 0 -u -z 1 -I control3cluster4r_r_cell.i2i -I c3c4F10_dres0_obj.i2i -S 2 1 0 c3c4F10.dpts
−
play new.i2i  -> ChainedTrans.txt
 
−
        register_images.pl ChainedTrans.txt
 
−
        dave -I new_trans.i2i -K -z 15  # a very big file.
 
−
 
 
−
source code: ~/krypton/bin/
 
−
see also:
 
−
combine use this on newer images which have spatial information stored in their header
 
−
parsepos.pl
 
−
~/krypton/Corvera/Olga
 
−
/storage/big1/lml/Corvera/Olga
 
−
recalc_quadrant.pl
 
−
recalc_quadrant2.pl
 
−
align_quadrants.pl the simplest manual method of aligning the four images in the well
 
  
 +
source for this cshell script is in /storage/big1/lml/was_invitro/unc/curvature
 
   
 
   
−
===  smooth_gaps.pl ===
+
</nowiki>
−
  This program applies smooth_gaps to all files named *reduced.i2i
+
===  find_vessels ===
−
  and produces files named *reduced_sg.i2i (unless -tag changed it).
+
  <nowiki>
  
−
  usage:
+
This program takes an image and scans it vertically in y at dx intervals.  It counts the number of
−
smooth_gaps.pl -run [options]
+
times a vessel of at least the specified width is entered.  A vessel is any pixel with the specified
 +
intensity value.  Each z slice in analyzed separately (and read in sequentially).  A count of vessels vs time and dx displacement is
 +
written to stdout.
 +
See /home/lml/krypton/Corvera/grant2010.  
  
−
  options:
+
Usage: find_vessels [options] image.i2i
−
-tag=something    tag to add before the .i2i in the new file name.
+
options:
−
-verbose   print out each filename as it is being smoothed.
+
  -z start stop: zplanes to analyze. 0-indexed inclusive. default is all z planes
−
-dummy   just echo what would be done, but don't do anything
+
  -i #: intensity of a pixel in the vessel. default = 32767
 +
  -w #: minimum vertical width of vessel in pixels, default = 2
 +
  -d #: dx step size, default = 100
 +
  -v: verbose
  
−
  see also:
+
Source in /home/lml/krypton/facil
−
smooth_gaps
+
See Also:
 +
Examples:
 +
 +
</nowiki>
 +
===  fit3gaussians  ===
 +
<nowiki>
 +
# /home/lml/krypton/bin/fit3gaussians -h
 +
This fits a 3 component (Gaussian diffusion) model to Joan Politz's data.
  
−
+
Usage:
−
===  trans.pl  ===
+
  fit3gaussians [options]
  
−
Takes a bunch of images taken in one well on the microscope, translates them
+
Description:
−
based on ChainedTrans.txt and combines them into one big image.
+
  Fits a three component Gaussian model to time series data.
  
−
ChainedTrans.txt should be a partially ordered file with entries like
+
Options:
−
X02:X03    (685,282):(736,287)  
+
  -a      : rather than having data fit a 3D diffusion model, have it fit a 2D model.
−
This means that location (685,282) in file X02 is the same as location (736,287) in file X03.
+
  -d # #  : only look at data sets from # to # (inclusive, first data set = 0).
−
By partially ordered I mean that for file1:file2 file1 should already have been the
+
            timings for the available data sets are (in seconds after uncaging):
−
right hand side of a similar line previously (so its position is already defined relative to
+
before_uncaging  0.050000  0.550000  1.050000  1.550000  11.550000  21.549999 
−
the file on the left hand side), unless it is the very first line in
+
31.549999  0.000000  0.000000  0.000000  0.000000  0.000000 
−
the file. So every right hand side is relative to a left hand side which has already
+
0.000000 
−
had its position defined.
+
            default data sets to look at are 1 to 4
−
Output goes to new_trans.i2i
+
  -e # #  : change the time in dataset # (0-indexed) to # (in seconds)
 +
  -f file : file name for test data (as produced by "distances" program).
 +
  -F file : file name for times for all the datasets from file, one time per line.
 +
            this also changes the default range to be all the sets
 +
            (use -d after this on the command line if you want to change the sets used).
 +
      note: there must be less than 1000 datasets.
 +
  -n # #  : number of function parameters and variances used for  
 +
            each parameter (default 10 paramters and  5 variances)
 +
  -p # #  : set function parameter number to some value (first # can be 0 to
 +
            the maximum number of parameters)
 +
  -s # # : set function paramter # start value to some value (first # can be 0 to
 +
            the maximum number of parameters)
 +
  -S # #  : set function paramter # variance value to some initial value (first # can be 0 to  
 +
            the maximum number of parameters)
 +
  -r # # #: set function paramter # range values to some initial values (first # can be 0 to
 +
            the maximum number of parameters)
 +
  -t #    : maximum number of random trials to search for parameters (default=100).
 +
  -T #    : search terminates when # successive times the smallest variance search is best. (default=5)
 +
  -m #    : maximum number of times to perform search, regardless of best variance (default=1000).
 +
  -M #    : maximum number of times try (for each parameter) to find a point within allowed range
 +
            default = 100
 +
  -D func #: print debug info. for specified function at level #
 +
              func 1: ars, levels: 1-4 with 4 being most info.
 +
              func 2: radial search, levels: 1-2
 +
              func 3: finding a test point, levels: 1-2
 +
  -P      :  just print out the data and theoretical curves for the start point and terminate.
  
−
register_images.pl [options] ChainedTrans.txt
+
example (see ~lml/vision/Politz/run)
−
-out=filename.i2i name of output file produced. default = newimage.i2i
+
default settings are:
−
-post=string the rest of the filename after what is in ChainedTrans.txt, eg, if the file is named
+
  t1 in seconds                  : range: -4.900E-02 to  5.500E+00, start: -2.500E-02
−
X01_0_0_1.i2i, -post=_0_0_1.i2i (this is also the default for post if not specified).
+
  D1 in microns^2/sec            : range:  1.000E-02 to  1.000E+02, start: 5.000E+00
−
-xdim=# size of original each original image (before recaling by maxtrima, see example below). default = 1392.
+
  mag1 in intensity/micron^3      : range:  5.000E+01 to  3.000E+04, start:  3.000E+03
−
-ydim=# default = 1040
+
  dc value                        : range:  0.000E+00 to  2.000E+03, start:  2.000E+02
−
-zdim=# default = 21
+
  D2                              : range:  1.000E-01 to  1.000E+03, start:  1.000E+02
−
-xscale=# binning done by matrixima (see example), coords will be scaled by this. default = 4.
+
  mag2                            : range:  5.000E+00 to  3.000E+04, start:  3.000E+02
−
-yscale=#
+
  t2                              : range: -4.900E-02 to  5.000E+03, start:  5.000E+02
−
-blend when combining images, blend the values where they overlap (default is to just replace any previous
+
  D3                              : range:  1.000E-01 to  1.000E+03, start:  1.000E+02
−
value with the new value).
+
  mag3                            : range:  5.000E+00 to  3.000E+04, start:  3.000E+02
−
    -dummy
+
  t3                              : range: -4.900E-02 to  5.000E+03, start:  5.000E+02
−
-debug
 
−
-h
 
  
 +
</nowiki>
 +
===  flood2zero  ===
 +
<nowiki>
  
−
-shift no circular shift (see tranima). old.
+
This program takes an original 3D image, a 2nd derivative image
−
note: tranima will pad images with -32768 which play will show in red (I think) and DAVE will be bothered by.
+
as produced by invariant -F 9 -b, and a list of 3d maxima and produces
 +
a new image which is just those voxels from orig.i2i that are negative
 +
(the 2nd derivative of a bright spot is < 0) in the derivative image
 +
and connected to a 3D maxima. So it is used in conjuction with invariant
 +
to identifiy bright blobs (eg, synapses) in a 3D image.
  
−
example:
+
Usage: flood2zero [options] orig.i2i deriv.i2i image.pts newimage.i2i
−
matrixima -row 4 W X Y Z -col 3 01 02 03 -pos 0 -obj 0 -fil 1 new.i2i
+
options:
−
play new.i2i  -> ChainedTrans.txt
+
  -b # : which resolution of maxima to use (object=noderiv_maxima# in pts file)
−
        register_images.pl ChainedTrans.txt
+
  -v #  : instead of using 0 as the object boundary cutoff, use #
−
        dave -I new_trans.i2i -K -z 15 # a very big file.
+
        -z low hi: zrange to examine, zero-indexed, inclusive.
 +
  -d dist: max chessboard distance (in voxels) from each point to flood fill.
 +
  -s size: maximum size in voxels of an object (will truncate to this size), default= 10000
 +
  -o file: save object (blob) statistics to specified file. This is very similar to
 +
what countobjs would produce when applied to newimage.i2i except newimage.i2i
 +
can have blobs which touch (due to the size contraints here) and hence show
 +
up as one large blob instead of two smaller blobs. Also, if the header for the
 +
image has a scale factor in it (which invariant writes) this scale factor will
 +
be used to convert all statistics back into original unscaled values.
 +
 
 +
note: entire image name (with extension) is NOT required.
 +
note: a - in place of an image name means stdin or stdout.
 +
note: a compressed image will be automatically be uncompressed.
 +
[source in /usr/people/lml/invitro/unc/curvature/flood2zero.c]
 +
 +
</nowiki>
 +
===  gsearch  ===
 +
<nowiki>
 +
program to do graph search on a slice of edge image
 +
USAGE:
 +
example gsearch -d name -x # -y # -a # -b # edge.i2i text.gpts
 +
    edge.i2i : magnitude input image
 +
    text.gpts: output is a graph points (gpts) text
 +
file having the same format as rpts file
 +
new boundary is stored under z=slice # (default: 1)
 +
 
 +
Options:
 +
-d  option specifies name of image containing direction of each edge
 +
    pixel of edge.i2i
 +
-k option specifies constant of proportionality
 +
    to be used with filgra (default: 1.0)
 +
-x  start x coordinate.
 +
-y  start y coordinate.
 +
-a  stop x coordinate.
 +
-b  stop y coordinate.
 +
-h prints this message.
  
−
source code: ~/krypton/bin/
 
−
see also:
 
−
combine use this on newer images which have spatial information stored in their header
 
−
parsepos.pl
 
−
~/krypton/Corvera/Olga
 
−
/storage/big1/lml/Corvera/Olga
 
−
recalc_quadrant.pl
 
−
recalc_quadrant2.pl
 
−
align_quadrants.pl the simplest manual method of aligning the four images in the well
 
  
 +
Source code in /storage/big1/lml/jac/was_invitro/graph
 
   
 
   
−
===  blur3d ===
+
</nowiki>
 +
===  histogram2 ===
 +
<nowiki>
 +
Usage:
 +
  histogram2 [options] boundary.i2i aimage.i2i bimage.i2i
 +
Description:
 +
  This program creates a number of histograms of intensity or volume as a function of
 +
  distance of the object voxels from the nearest background voxel.
 +
  Requires:
 +
    boundary.i2i - boundary image created with boundary or createBoundary.
 +
object voxels are those with a value of 6
 +
background voxels are those with a value of 4
 +
border voxels are those with a value of 2
 +
all other voxels are ignored.
 +
            NOTE: by default this is assumed to be padded by 7 pixels all around.
 +
                  ie, IT IS LARGER THAN aimage.i2i and bimage.i2i
 +
                  createBoundary does this. see -p if you don't want this default.
 +
    aimage.i2i  - data image a.
 +
    bimage.i2i  - data image b.
 +
  Returns:
 +
    aimage_H.histo - output file with histogram info (-H can change this).
 +
  Output file format (10 columns):
 +
  1. bin number (i.e., distance from background) (each bin = 0.1 pixels)
 +
      note:a pixel touching the boundary is a distance of .5 (center is .5 from edge)
 +
  2. histogram of intensities in image a.
 +
  3. number of pixels in this bin (i.e., volume at this distance).
 +
  4. histogram of intensities in image b.
 +
  5. histogram of intensities in image a > athreshold (intens-thresh if -s option used).
 +
  6. histogram of intensities in image b > bthreshold (intens-thresh if -s option used).
 +
  7. histogram of intensities in image a, where values in a are such that
 +
      if (image a > athreshold)  and (image b > bthreshold), a=a (a-athresh if -s option) else a=0.
 +
  8. histogram of intensities in image b, where values in b are such that
 +
      if (image a > athreshold)  and (image b > bthreshold), b=b (b-bthresh if -s option) else b=0.
 +
  9. # of pixels in image a > image a threshold (at this distance), like 3 but only
 +
      counting voxels > threshold.
 +
  10.# of pixels in image b > image b threshold (at this distance), like 3 but only
 +
      counting voxels > threshold.
 +
 
 +
Input Options:
 +
  -o    :  border pixels are background voxels (default is object voxel)
 +
  -p x y z  :padding in x,y,z included in the boundary image  (default 7 in all directions).
 +
          (boundary image found in the programs boundary and
 +
            createBoundary are by default padded by 7 pixels
 +
            on all sides of the image.
 +
 
 +
Distance Options:
 +
  -P x y z : don't find histogram of all pixels to boundary, just the distance
 +
            of this pixel from the boundary. Can be used multiple times on command line.
 +
            (x,y,z) is 1-indexed (like play) aimage.i2i coordinate values.
 +
  -d #:    maximum distance (in pixels) to check, default = .5*(min dim)
 +
          Any pixel further than the maximum distance is considered to be at the maximum distance.
 +
          This apply also for the -ta and -tb distance based thresholds.
 +
  -x #  :  set x weight to # (default 1.0).
 +
  -y #  :  set y weight to # (default 1.0).
 +
  -z #  :  set z weight to # (default 1.0).
 +
  -F    :  Fast histogram - voxel level (default fast).
 +
  -S    :  Sub-voxel resolution histogram(default fast).
 +
  -Y # #:  limit Y direction from # to # (default off).
 +
          If the long axis of the cell is in the in the y direction,
 +
          and the cell is cut off, using the -Y option will limit searching
 +
          for the boundary in that direction, i.e. the ends past these values will
 +
          not be considered part of the boundary.
 +
          (similarly true for -X and -Z options)
 +
  -X # #:  limit X direction from # to # (default off), inclusive, 1-indexed.
 +
  -Z # #:  limit Z direction from # to # (default off).
 +
  -c:    pixels outside the image are considered border pixels.  For example, if the outlines
 +
          used to create boundary.i2i were in z=1, but you want pixels inside the z=1 boundary
 +
          to be counted as being 1 pixel away from the boundary (eg, from the phantom z=0 slice)
 +
          use this option.
 +
 
 +
Threshold Options:
 +
  -a #  :  set aimage threshold to # (default 0.000000).
 +
  -b #  :  set bimage threshold to # (default 0.000000).
 +
  -s :    subtract thresholds (and then clamp to >=0) rather than simply keeping intensities > threshold.
 +
          This is more like what you might want to do to subtract off background or nonspecific flourescence.
 +
  -ta athresh.txt  :  apply different thresholds at different distances to aimage.i2i.
 +
                    athresh.txt as produced by analyze_histo (applied to -f aimage_H.i2i output)
 +
  (i.e., 2 columns: bin # (tenths of a pixel) and threshold).
 +
  -tb bthresh.txt  :  like -ta, but for bimage.i2i
 +
  -ra # :  rescale athresh.txt prior to use, multiply values in file by #.
 +
  -rb # :  rescale bthresh.txt prior to use, multiply values in file by #.
 +
 
 +
Ouput Options:
 +
  -f aimage_H.i2i xdim ydim:  print out a full histogram of aimage. This is a
 +
          2D image (eg, aimage_H.i2i) with the specified x and y dimensions.
 +
          The x direction is pixel intensity, and the y direction bin number (tenths of a pixel) and
 +
          the pixel value wll be set to the number of voxels at that distance and that intensity.
 +
          Specify -1 for xdim or ydim if you want that dimension calculated automatically
 +
          (xdim will be approx. max intensity in image and ydim about 10*max dist, see -d option).
 +
          Explicitly set the dimensions if you will want to add this image with others (since they must all
 +
          be the same dimensions), e.g., to average data from a set of control files.
 +
          note: -S option will cause the "number of voxels" to appear to change.
 +
          Use the analyze_histo program to produce a distance dependent threshold file from this (e.g. to read
 +
          back into histogram2 on another pass via the -ta or -tb option).  This image can be large (25MB).
 +
  -H file: Output filename for histogram (instead of aimage_H.histo).
 +
  -A aimage_thresh.i2i  :  write out aimage.i2i after threshold is applied (for visualization purposes)
 +
                          these are the pixel values used to produce column 5 of aimage_H.histo
 +
  -B bimage_thresh.i2i  :  write out bimage.i2i after threshold is applied (for visualization purposes)
 +
                          these are the pixel values used to produce column 6 of aimage_H.histo
 +
 
 +
Miscellaneous Options:
 +
  -D distimage.i2i  :  print the distance image (for debugging). Each voxel is 100 times its distance
 +
    from the membrane (a pixel neighboring the background has a distance of .5 pixels).
 +
                      This can also be used by mask_image to then mask desired pixels.
 +
  -h    :  print this message.
 +
 
 +
Notes:
 +
  All the images should be oriented in the same direction.
 +
  Entire image name (with extension) is required.
 +
  A - in place of an image name means stdin or stdout.
 +
 
 +
 
 +
Example:
 +
  create_dcim -d 4 -s 234 294 24 -| addlines -P -1 - rob15_nucleus.rpts -|
 +
            flood -e -1 -E 2 -I 6 -b 4 - mask.i2i
 +
  histogram2 -p 0 0 0 -Z 2 23 -d 40 -a 3552 -b 7767 -z 1.34 mask.i2i rob15f_rtm.i2i rob15r_rtm.i2i
 +
 
 +
Example: use a variable distance threshold to subtract background flourescence from dual labeled images:
 +
    addlines -P -1 control_aimage.i2i control_membrane.rpts -| flood -e -1 -E 2 -I 6 -b 4 - mask.i2i
 +
        # create a full histogram so we can use a variable threshold based upon distance from membrane:
 +
    histogram2 -p 0 0 0 -d 40 -z 1.34 -f controla_H.i2i mask.i2i control_aimage.i2i control_aimage.i2i
 +
        # create the variable threshold file:
 +
    analyze_histo controla_H.i2i > controla_H.thresholds
 +
        # repeat for control bimage (e.g., second wavelength):
 +
    histogram2 -p 0 0 0 -d 40 -z 1.34 -f controlb_H.i2i mask.i2i control_bimage.i2i control_bimage.i2i
 +
    analyze_histo controlb_H.i2i > controlb_H.thresholds
 +
        # now apply these thresholds to the real dual labeled (not the control) data:
 +
    addlines -P -1 real_aimage.i2i real_membrane.rpts -| flood -e -1 -E 2 -I 6 -b 4 - real_mask.i2i
 +
    histogram2 -p 0 0 0 -d 40 -z 1.34 -s -ta controla_H.thresholds -tb controlb_H.thresholds \
 +
                    real_mask.i2i real_aimage.i2i real_bimage.i2i
 +
See Also:
 +
  boundary, createBoundary, analysis, analyze_histo, threshold, nucleus_overlaps, mask_image, Corecell.pl.
 +
 
 +
Source code in /storage/big1/lml/jac/was_invitro/Projects/boundary/boundary
 +
 
 +
 
 +
</nowiki>
 +
===  image2ptsL  ===
 +
<nowiki>
  
−
Blurs an image by a psf.  
+
This program takes an image, and identifies boundary pixels of objects.
−
FFTs are done with 4 byte floats, so total space needed will be 4 times the padded image size (in voxels)
+
Boundary pixels are foreground pixels which have at least one
−
plus the size of the two original images (image and psf)
+
backgound pixel among their 8 neighbors.
−
The input image can be real or short int format. The psf must be short int format.
+
In addition, any foreground pixel which is on the edge of a 2D
−
note: performs a convolution (not a correlation).
+
slice is also considered a boundary pixel.
−
note: fixed a small bug. 10/17/11
+
These boundary pixels are written out in "addlines" format.
 +
Each zslice is analyzed sequentially.
 +
Points are unordered, and they are all considered part of one object.
 +
Points in the pts file have their value set to -1
  
−
Usage: blur3d [options] image psf newimage
+
Usage: image2pts [options] image.i2i pointfile.pts
 
options:
 
options:
−
  -S #:  scale (multiply) output values by # (to prevent underflow or overflow)
+
  -t #: threshold, above which is foreground. Default = 0
−
  voxels which after scaling are > 32767 are set to 32767. voxels < -32766 are set to -32766.
+
note: entire image name (with extension) is NOT required.
−
    -p # # #: pad the image in x, y, and z by the specified amounts
+
note: a - in place of an image name means stdin or stdout.
−
by default image is padded (with 0) by 1/4 psf width all around,
+
note: a compressed image will be automatically be uncompressed.
−
so its size for blurring becomes bigger by psf/2.
+
−
psf is always padded to the same size the image becomes.
+
</nowiki>
−
Padding is stripped off before newimage is written, so it will be the same size as
+
===  invariant  ===
−
the original image.
+
  <nowiki>
−
    -N # # #: pad so that the image has the specified dimensions after padding (just prior to FFT).
+
 
−
  -n #: pad with # instead of boundary pixel values.
+
This program takes an image, blurs it with a Gaussian of
−
  -Z:   pad with zero instead of the boundary pixel values.
+
specified standard deviation and then takes derivatives of the image
−
  -c # # #: the center of the psf is at (#,#,#) zero indexed.  
+
It does all of this in the frequency domain.
−
                the default is to take the brightest pixel as its center.
+
 
−
  -C:  don't center the psf at all (default is to center on the max, unless -c is specified).
+
outfile is the root name for derivative of blurred images (outfile_dres#.i2i, unless -M),  plain blurred
−
  -P:   normalize the psf so each slice (after padding) averages to 1. -P acts like an optical PSF.
+
images (-b option, outfile_bres#.i2i) and/or pts files (-m or -M options, outfile.pts).
−
        the default is to normalize so the entire psf (after padding) has an average pixel value of 1
+
 
−
              (the default keeps total light constant).
+
Any function which is a ratio of sum of products of derivatives (eg, (LxxLy + LyzLx)/(LyLy+LzLz))
−
  -d:   don't normalize at all.
+
can be calculated.  Edit file init_invariant.c in
−
  -o:   do a cross correlation instead of a blur (convolution)
+
/usr/people/lml/invitro/unc/curvature and recompile on molmed if you want a function which
−
  -R:   output image as 4 byte floating point instead of short integer.
+
is not listed here.
−
  -v:  verbose. print more info to stderr.
+
 
 +
Usage: invariant [options] infile.i2i outfile
 +
options:
 +
  -F #:  function to calculate. Default = 1. Available functions are:
 +
  -F 1: LxLx+LyLy+LzLz : 3D gradient squared
 +
  -F 2: .5(Lxx+Lyy+Lzz)/sqrt(LxLx+LyLy+LzLz) : normalized 3D Laplacian
 +
  -F 3: LxLx+LyLy : 2D gradient squared
 +
  -F 4: sqrt(LxLx+LyLy) : 2D gradient
 +
  -F 5: 2(LxxLyy-Lxy^2)/(Lxx^2 + Lyy^2 + 2Lxy^2): 2D light or dark blobs (p. 252 IPMI '91)
 +
  -F 6: 2(LxxLyy-Lxy^2), same a -F 5, no normalization (so noise isn't magnified).
 +
  -F 7: mean curvature of 3D isointensity surface normalized by gradient squared
 +
  -F 8: mean curvature of 3D isointensity surface, no normalization.
 +
  -F 9: 2nd der. in the grad. direction times gradient squared.
 +
  -F 10: 2nd der. in the grad. direction (should = 0 at locations of edges)(use -u option).
 +
  -F 11 XYZ    e.g., -F 11 021  take zero derivatives in X direction, 2 in Y, 1 in Z (Lyyz).
 +
by calling invariant multiple times with various XYZ and using the -g option you
 +
can create a bunch of derivative images which you can then read in and combine
 +
however you like (e.g., my curvature_eigenvectors program)
 +
  -N #: normalize image intensities prior to use to be between 0 and #
 +
this performs: newval = #*(val-minval)/(maxval-minval)
 +
  -u #: during division (-F 2,5,7,10) set divide by 0 to this integer, default=0.
 +
        note: this is the final output pixel value, after any scale factors (-r or -R).
 +
        a good value is -32768, the minimum possible output pixel value.
 +
  -s sigx sigy sigz:   standard deviation in spatial domain of Gaussian.
 +
  -S sigx sigy sigz:   standard deviation in frequency domain of Gaussian.
 +
default = 3.402823E+38  3.402823E+38  3.402823E+38
 +
  -l # res2 res3 ... :  # = number of different resolution levels to calculate.
 +
res2 ... res# are the relative sigmas for each resolution level after
 +
the first one. eg, -l 3 .9 .7 means that 3 blurred levels should
 +
be calculated and the sigma (in the freq domain) of the second
 +
level should be .9 of the first level (ie, more blurred), and the 3rd
 +
level should be .7 of the first level (as specified with -S).
 +
  -L # factor:  # = number of resolution levels. factor is the relative sigmas in
 +
the spatial domain for each level relative to the previous level.
 +
  -e epsilon:  cutoff for Gaussian intensity calculation.
 +
default = 0.040000
 +
  -r: apply scalefactor to output so min= -32766 or max= 32766. This is useful
 +
especially when all the values would otherwise be < 1.
 +
This also prints to stderr the min and max values before rescaling.
 +
  -R #: rescale output by multiplying all values by #.
 +
  -m: calculate coords of 3D maxima after blurring and taking derivatives.
 +
Creates an outfile.pts file.
 +
  -M: same as -m, but also prevents writing out blurred images.
 +
        -b:         calculate coords of 3D maxima after blurring (but before derivatives). This
 +
        can be used with the -m option. Objects associated with this option will
 +
        be named "noderiv_maxima#", where # is the resolution number.
 +
        This is useful for passing to flood2zero to find the extent of the bright spots.
 +
Also writes out blurred images (outfile_bres#.i2i).
 +
  -B: same as -b, but prevents writing out the blurred images.
 +
  -n #: maxima must have an intensity > # (must use -m, -M, or -b with this)
 +
  -f #: max pixel must be > all neighbors by at least # to be a maxima. default = 0.000100
 +
  -g: output a floating point image (no rescaling done). This output can be used
 +
by my curvature_eigenvectors program. -u option still applies.
  
−
note: entire image name (with extension) is NOT required.
+
note: image dimensions which factor to 2, 3, or 5 go fastest with the FFT.
−
note: can read from stdin or write to stdout (use a dash instead of the file name)
+
note: the entire image name is NOT required.
−
note: a compressed image will be automatically be uncompressed.
+
note: compressed images will be uncompressed.
 +
note: a - in place of an image name means stdin or stdout.
 +
see also: max3d, multi_gauss_deriv, flood2zero, find_blobs, blur_spatial, canny, zerosu.
 +
source in /storage/big1/lml/was_invitro/unc/curvature
 +
/home/lml/krypton/bin/invariant
  
−
Copyright 2010 University of Massachusetts Medical School and the Biomedical Imaging Group
+
</nowiki>
−
All rights reserved. Explicit permission to use this program must be received prior to use.
+
===  linux_epr_x86_64-7-12-2012  ===
 +
<nowiki>
 
   
 
   
−
===  canny  ===
 
  
−
This program reads in a 2 or 3d image and outputs the results
+
epr                              UMMC/BIG                          Jul 12 2012
−
of a 2D Canny edge detector on it. Canny is the derivative
+
 
−
(in the gradient direction) of the magnitude of the first
+
NAME
−
derivative (so it is a type of 2nd derivative).   
+
    epr - Exhaustive Photon Replacement (EPR) restores contrast by removing
−
NOTE: see option -n below for an exact answer.
+
    residual out-of-focus light and improves resolution while maintaining
−
Each z slice is treated
+
    numerical accuracy of 3-D images of specimens obtained with serial
−
independently of the others. Its application will often be
+
    optical sectioning from wide-field or confocal light microscopy.
−
preceded by a Gaussian blur and followed by the calculation
+
 
−
of zero crossings.
+
  SYNOPSIS
−
If an INT type image is input, an INT is output.  
+
    epr [options] before-image[.tif] after-image[.tif]
−
If a REAL type image is input, a REAL is output.
+
 
 +
DESCRIPTION
 +
    epr performs regularized, iterative image restoration with a non-
 +
    negativity constraint.  The before-image is a three dimensional (3-D)
 +
    TIFF image composed of rectangular, regularly spaced, optical sections.
 +
    Large images are decomposed into smaller, overlapping (in x and y only)
 +
    image segments for restoration, and the restored segments are recomposed.
 +
    The after-image is the restored 3-D image. All options must appear
 +
    before the image file names, but the order of options is not important.
  
−
Usage: canny [options] infile outfile
+
    epr has the following options:
−
  options:
 
−
-d: Debugging flag
 
−
-n: normalize the result. This option causes the true 2nd
 
−
derivative to be calculated. Without it, only the sign
 
−
of the second derivative is guaranteed to be correct
 
−
(which is sufficient if only zero crossings will be
 
−
calculated from it). Divides each point by Ix*Ix+Iy*Iy.
 
−
-s #:  scale factor to multiply all output values by. This is
 
−
useful if you are outputing an INTEGER image with a very
 
−
small range of values which would otherwise all be truncated
 
−
to just a few numbers.
 
−
note: entire image name (with extension) is NOT required.
 
−
note: a - in place of an image name means stdin or stdout.
 
−
Example usage:
 
−
To create a gradient edge strength image for a shrink wrap run:
 
−
  blur_spatial image.i2i -|canny - -|zerosu - - |blur_spatial - out.i2i
 
−
 
−
===  cblur  ===
 
−
 
−
Usage: cblur [options] arg1.i2i arg2.i2i
 
−
  For every zero valued point in each xz plane,  cblur
 
−
  replaces the zero value with the -100 times the distance
 
−
  from the nearest non-zero voxel in the xz plane.
 
−
Options: None at this time.
 
  
−
Note: entire image name (with extension) is required.
+
    -h                Prints basic help on syntax and available options.
−
Note: a - in place of an image name means stdin or stdout.
 
  
 +
                              Required Parameters
  
−
=== center_of_mass  ===
+
    -psf file[.tif]  The 3-D point spread function image file. The psf must
 +
                      match the optical configuration used for the before-
 +
                      image. The psf depth (number of z-planes) need only be
 +
                      less-than or equal-to twice that of the before-image.
 +
                      Extra z-planes are symmetrically discarded.
  
−
This program reads in a 4D image and translates each 3D image
+
    -smoothness alpha The alpha value where alpha is typically 0 < alpha < RNL.
−
so its center of mass is moved to be the center of the image.
+
    -sm alpha        RNL is the residuals noise limit as reported by prepdata
−
One 3D image is read at a time.
+
                      (see also). Smaller values of a correspond to less
 +
                      smoothing. (RNL)^2 is usually a good starting choice for
 +
                      alpha.
  
−
Usage: center_of_mass [options] inimage.i2i outimage.i2i
+
    -iterations n    Maximum number of iterations to perform. Iterating may
−
options:
+
    -it n            terminate earlier if convergence is detected. (see
−
  -t tdim:  number of time points, default = 1. This is NOT read from the header.
+
                      -convergence)
−
  -T #:  threshold inimage.i2i prior to analysis. voxels <= # ignored, default = 0
+
 
−
  -v: verbose
+
                              Control Parameters
 +
 
 +
    -scaling n        where n is after image scaling factor. Used to prevent
 +
    -sc n            integer overflow when saving after image to file.
 +
                      Default=1.0
  
−
note: source code in krypton:/home/lml/krypton/facil directory.
+
    -convergence n    Criteria for terminating iteration, where n is << 1.
−
+
    -co n            0.00001 represents true convergence. 0.001 usually
−
===  combine_images  ===
+
                      achieves 90-95% convergence in about half the number
−
This program reads a bunch of image files which have their xy position within a well stored in their header.
+
                      of iterations. Values of n larger than 0.001 are not
−
It then combines the images. Karls acquisition program for the Zeiss microscope should automatically add
+
                      generally recommended. Default=0.001
−
this information to the header. A maximum of 50 images can be combined. Images must all be the same size.
 
−
Images are also assumed to be aligned in z already. Images are added to the output image in the order specified
 
−
on the command line, with later pixel values replacing previous values if there is an overlap of the images.
 
  
−
Usage: combine_images [options] im1.i2i im2.i2i [more input images]  outim.i2i
+
    -axial n1 n2      The z-axis object extrapolation beyond the sectioned
−
options:
+
    -ax n1 n2        image data where n1 is before first plane and n2 is
−
-d debug. stuff to stderr.
+
                      after last plane. n is in planes.  By default,
−
-f do NOT fill pixel locations which have no data with nearest data value (added 2/3/14)
+
                      extrapolation extends +-1/2 the z-axis extent of the point
−
Filling is done by default to try to avoid introducing sharp intensity edges into the image.
+
                      spread function image and should be sufficient. Smaller
−
-X flip each input image in X before using it (-Y can also be specified at the same time)
+
                      values of n1 or n2 have the effect of spatially
−
-Y flip each input image in Y before using it
+
                      constraining the restored object and should be applied
−
-p # over-ride the specified pixel size with this one (in um). The 2.5X objective has a
+
                      carefully.
−
pixel size of 2.58 um stored in the image header (for images acquired prior to 4/13/13)
 
−
but it should be 2.41
 
−
-x #:#[:#...]  rather than use the x positions stored in the image header use these x pixel positions.
 
−
these should be the x offset of one image relative to another, in the same order as the
 
−
image names on the command line (there MUST be one number for each image). For example:
 
−
combine_images -x=0:500:0:500 im11.i2i im21.i2i im12.i2i im22.i2i outim.i2i  if im21.i2i and im22.i2i
 
−
are displaced 500 pixels to the right of the other two images.
 
−
-y #:#[:#...]  same, but for displacement in the y direction. Since these are pixels, they should be integers.
 
−
note: many times images are 1392x1040 and overlap by 50 pixels, and image 0 is bottom left
 
−
      image 1 is top left, image 2 is bottom right, and image3 is top right, then:
 
−
      -x 0:0:1342:1342 -y 0:990:0:990  should work
 
−
-o # rather than specifying -x and -y, thus just specifies the overlap (e.g., 50) and figures out
 
−
the rest for you (assumes images ordered BL,TL,BR,TP - as above - and x and y overlap the same
 
−
works for any size image.
 
  
−
note: entire image name (with extension) is NOT required.
+
    -time n1          The timepoint where you want the restoration to occur (0 indexed)
−
note: a - in place of an image name means stdin or stdout.
+
    -t n1
−
note: compressed images are temporarily uncompressed into /usr/tmp.
 
  
−
See also: parsepos.pl (not needed now that Karl fills the image header with this info)
+
    -threads n        The number of threads FFTW should use. Default=1
−
    combine_images.pl    applies combine_images (and, optionally, reduceima) to all files in a directory.
+
    -th n
−
    register_images.pl
+
 
−
    align_well_images.pl    pre-processor for combine_images if the y-coord in the header is invalid.
+
    -transverse n    The x-axis and y-axis object extrapolation in pixels.
−
    insert_image
+
    -tr n            The epr process extrapolates the restoration beyond the
 +
                      bounds of each image segment in order to account for
 +
                      exterior out-of-focus contributions. Default = 1/4 of X or Y
 +
                      dimsenion PSF width, which ever is larger.
 +
 
 +
EXAMPLES
 +
    To see the command line syntax
  
−
Source code in ~/krypton/facil/combine_images.c
+
      % epr
−
combine_images:
+
    or
 +
      % epr -h
  
−
== Miscellaneous ==
+
    Given specimen image mycell_.i2i and point spread function image mypsf_.i2i
−
===  analyze_cross_sections_of_voronoi.pl  ===
+
      let smoothness equal 0.0005, maximum number of iteration equal 250
 +
      and convergence equal 0.001. Perform single-resoltion(default) EPR:
  
−
This program reads files produced by cross_sections_of_voronoi (-P)
+
      % epr -psf mypsf_ -smoothness 0.0005 -iterations 250 mycell_ mycell_r
−
Results prints volume (x) vs predicted volume (y) to stdout.
 
  
−
analyze_cross_sections_of_voronoi.pl [options] file1.txt [... filen.txt]
+
FAQ (Frequently Asked Questions)
  
−
options:
+
      see  http://...
−
-debug info about all fields on each line printed to stderr
+
FILES
−
-stdout everything to stdout
+
        xcomp:/home/epr/epr
−
-print_avg for each file, just graph mean and std error of the mean, rather than all data pts
+
        xcomp:/home/epr/cs_epr.lo
  
 +
SEE ALSO
 +
        prepdata
 +
        preppsf
  
−
examples:
 
−
cross_sections_of_voronoi -P voronoi.i2i > filename1.txt
 
−
cross_sections_of_voronoi -P voronoi.i2i > filename2.txt
 
−
        axnalyze_cross_sections_of_voronoi.pl *.txt
 
  
−
see also:
+
</nowiki>
−
/storage/big1/lml/Raz/voronoi/README, true_vs_predicted.xmgr
+
===  many_overlapsL  ===
−
/home/lml/krypton/Corvera/Raz/stereology/README, run_voronoi.pl, run_voronoi2.pl, compare_volume_measurements.pl, plot_results.pl
+
<nowiki>
−
cross_sections_of_voronoi
+
insufficient number of arguments
−
/home/lml/krypton/packages/fiji/Fiji.app/macros/stereology_lml.txt
 
−
/home/lml/krypton/packages/fiji/run_stereology.txt
 
−
/storage/big1/lml/Raz/voronoi/
 
  
 +
This shell script calls overlap many times for a range of thresholds
 +
in both images.
 +
Unknown options are passed to overlap.
 +
Output is put into the specified output file
 +
This should then be processed by the program: create_images
 +
Note: a temporary file is created in the current directory.
 +
See also: create_images, overlap, probcolocal
 
   
 
   
−
===  neartrack.pl ===
+
USAGE: many_overlaps [options] image1.i2i image2.i2i output
−
neartrack.pl
+
options:
 +
  -t low hi step :range of thresholds for image1.i2i
 +
default = 1, 101, 10
 +
  -T low hi step :range of thresholds for image2.i2i
 +
default = 1, 101, 10
 +
 +
</nowiki>
 +
===  map_to_surface ===
 +
<nowiki>
 +
error: 
 +
There should be 3 or 5 main arguments (exluding options).
 +
Arguments seen:
 +
This program takes a surface (which are those voxels which are nonzero)
 +
and one (or two) data images.  It then maps (moves) the data voxels (nonzero
 +
voxels) to the position of the nearest surface voxel.  When done, one (or two) new
 +
image is produced which has the data values at these new positions.
 +
In addition, some statistics about the mapping are printed to stderr.
  
−
  find which track a specified coordinate is near (ie, with dx,dy) (eg, DAVE crosshairs gives us a coord, what track is it?)
+
NOTE:A surface can be generated by Simple_2D_wrap or shrink_wrap. This "mesh"
−
  assumes coords passed are zero indexed
+
representation of the surface (which DAVE can display using the -i option)
 +
can be converted into a surface .i2i image for map_to_surface using
 +
/usr/people/jac/cell/bin/make_surface -I surface.i2i -c 1 mesh.i2i surface.rpts
 +
The resulting surface is made up of voxels. Voxels which are just below the
 +
surface will be considered to be a distance of 1 voxel from the surface
 +
(the distance between the centers of the two voxels).
  
−
    usage:  neartrack.pl -x=# -y=# [-z=#] [-dx=#] [-dy=#] [-oneindexed] file.rpts
+
Usage:
 +
   map_to_surface [options] surface.i2i image.i2i out_image.i2i [image1.i2i out_image1.i2i]
 +
Options:
  
−
+
a second input image and output image can be specified on the command line.
−
===  parse_macros4.pl  ===
 
−
Can't use 'defined(%hash)' (Maybe you should just omit the defined()?) at /home/lml/krypton/bin/parse_macros4.pl line 562.
 
  
−
=== path_diff.pl ===
+
-c: rather than mapping data values to the nearest surface voxel, map
−
compare the differences between the paths found from two different runs of exo5
+
a count of the number of data values which mapped to the
 +
surface voxel. I.e., if 2 data voxels mapped to a surface voxel, a
 +
2 is stored there (rather than the max of the 2 data values-the default).
 +
-b: rather than mapping data values to the nearest surface voxel, or
 +
a count, just map a 1 (ie, binary) onto the surface where ever a data
 +
voxel maps (regardless of how many map there or their intensity).
 +
-m: surface.i2i is actually a mesh file, not an image file
 +
produced from a mesh file. In this case, only map data
 +
values to surface vertices (since map_to_surface doesn't
 +
yet know how to convert mesh files to complete surfaces
 +
like make_surface).
 +
      -d #:  minimum distance (in voxels) which data must be from the surface and
 +
still be mapped onto it.  Data closer than this is ignored.
 +
By default all data is examined (ie, min_distance = 0)
 +
      -D #:  maximum distance (in voxels) which data can be from the surface and
 +
still be mapped onto it.  Data further than this is ignored.
 +
By default all data is examined (ie, max_distance = 1/2 the minimum
 +
image dimension, this should include all of the data).
 +
      -x:    exclude data voxels which are already on the surface (they will be zeroed).
 +
(note: setting the minimum distance > 0 will not achieve the same effect since
 +
that would still allow a data voxel on the surface to match to a surface
 +
voxel at a neighboring position).
  
−
path_diff.pl file1 file2
+
Source code in /storage/big1/lml/was_invitro/Moore/pkc
 +
 
 +
</nowiki>
 +
===  min_cylinder  ===
 +
<nowiki>
 +
MIN_CYLINDER program - Version 1.0
 +
 
 +
This program takes an initial cylindrical mesh surface
 +
and allows it to deform to minimize a function of
 +
surface curvature and stretch and image intensity
 +
 
 +
Usage: min_cylinder [options] insurface infile outsurface
 +
 
 +
OPTIONS CONTROLLING TERMINATION CRITERIA:
 +
[-n #] Maximum iterations
 +
[-c #] Minimum pixel changes per iteration
 +
 
 +
OPTIONS CONTROLLING SURFACE SHAPE:
 +
[-w # # # # # #] Weights for Su Sv Suu Suv Svv E
 +
default = 0.20, 0.20, 0.10, 0.20, 0.10, -0.10
 +
NOTE: for a 2D time series, the u
 +
direction is usually in the xy plane
 +
and v is the time direction.
 +
-b:  use bilinear interpolation along the u coordinate
 +
      direction to calculate the energy component due to
 +
      the data. This option only works in conjunction with
 +
      the -D option. This causes all data along the boundary
 +
      in each xy plane to contribute to the energy of the
 +
      surface, instead of just voxels at the locations of the
 +
      vertices of the surface.
 +
      NOTE: does not use weights specified by -W option yet.
 +
-a:  same as -b but the average energy along the boundary is
 +
      calculated instead of total, so long lines are not preferred.
 +
-p:  same as -a but the average energy along each boundary segment
 +
      is weighted by the percentage of total boundary length which
 +
that segment contributes. So long bad lines count more than
 +
short bad lines.
 +
-g:  weight the data along the boundary by the dot product with the
 +
unit length vector normal to the gradient. Thus boundary segments which
 +
are parallel to the data gradient direction will count less than
 +
they otherwise would. Use with -b, -a, or -p option.
 +
-z #: the cost of a zero length line (before applying w6)
 +
      default = 0, only valid with -a or -b option.
 +
-N: use "normal" curvature instead of parameterization
 +
dependent curvature calculation.  This option should produce
 +
a more correct curvature calculation, so the program does not
 +
get fooled into thinking that small objects have smaller
 +
curvature.
 +
 
 +
OPTIONS CONTROLLING ALLOWABLE SURFACE MOVEMENT:
 +
[-D]  Restrict movement to 2 dimensions (x & y)
 +
[-s]  Skip first and last planes of cylinder
 +
      (so they won't move at all).
 +
[-S filename] filename of z planes to skip (do not use with -s).
 +
[-f]  Fix first and last planes of cylinder to move only in x&y
 +
[-F filename] filename of z planes to fix to move only in x&y
 +
(do not use with the -f option).
 +
[-A]  when the -F or -S flag specified, normally if two adjacent
 +
      slices are specified in the rpts file then the surfaces on
 +
      either side of these become independent of the each other.
 +
      If -A is specified then they will still affect each other.
 +
 
 +
OPTIONS CONTROLLING INPUT, OUTPUT, AND DEBUGGING INFO:
 +
[-W # # #] Weights for scaling x y z dimensions.
 +
[-r rescale]  rescale image data from 0 to rescale
 +
before using. default = 1000.000000
 +
    If rescale <0 then no rescaling will be done.
 +
[-u]  expect input image to be i2i - output also i2i.
 +
[-I #] write out vertex coords every # images
 +
      written in unc image format. can be read
 +
      in and displayed by wire. If SAVE_ENERGY is
 +
  defined then energy of each vertex also saved.
 +
      Currently SAVE_ENERGY is defined.
 +
[-i]  Copy information fields flag
 +
[-t "title"]  New image title
 +
[-d #]  Debugging level, 1 (least) to 2 (most)
 +
 
 +
See also: Simple_2D_Wrap, 2D_wrap, shrink_wrap
 +
 
 +
</nowiki>
 +
===  morph3d  ===
 +
<nowiki>
 +
 
 +
This program does morphological processing on greyscale images. Currently the
 +
default intensity it filters with is a constant, although different shapes and
 +
intensity profiles are allowed.  Think of a 2d image as a 3d binary array of voxels
 +
where the height dimension is the intensity of the image at this x,y position.
 +
All those pixels which are less than or equal to the height are 1, others are zero.
 +
This object (made up of voxels which are 1) is the UMBRA of the image.
 +
 
 +
Dilation: Take the umbra of
 +
the image and the umbra of the filter. Then the image dilated by the filter is
 +
those voxels which get "reached" by the filter voxels when the filter umbra
 +
is translated through the image umbra (i.e., move the origin of the filter umbra
 +
everywhere inside the image umbra, all those voxels ever reached by the filter are
 +
part of the dilated umbra).
 +
For a constant intensity filter this reduces to just taking the
 +
maximum of the image intensities at the filter's locations.
 +
 
 +
Erosion: similar to dilation, but now as you move the filter around, only keep
 +
those voxels which, when the filter umbra is centered on them, permit all the
 +
filter voxels to lie on top of image voxels (i.e., the filter umbra fits inside
 +
the image umbra when centered on that voxel).Erosion is dilation of the background.
 +
This reduces to a minimum for a constant intensity filter.
 +
 
 +
Opening: erosion followed by dilation, can be visualized by taking the filter umbra
 +
surface and pushing it hard up under the image umbra surface. The highest voxels
 +
reached by the filter surface represent the new filtered image surface.
 +
(e.g. the "solvent or probe accessible surface" from below.
 +
 
 +
Closing: dilation followed by erosion.  Dilation can be visualized as
 +
flipping the filter umbra and rolling it along the top (outside) of
 +
the image umbra and recording the lowest voxels reached. (e.g.,the solvent
 +
accessible surface from above.).
 +
 
 +
If a 3D image is input, and a 3D filter shape is specified, then all analysis
 +
is done using 3D morphological filtering.  If a 2D filter is specified, then each
 +
zplane is analyzed separately.
 +
 
 +
Usage: morph3d [options] inimage.i2i outimage.i2i
 +
options:
 +
  -v: verbose, prints out which z slice is being analyzed to stderr.
 +
  -b #: any pixels <= # are background, ignore them. default = -32766
 +
  -h #: the maximum height (intensity) of the filter center relative to other
 +
                      entries in the filter. The default =0, ie all entries in the
 +
                      filter are at the same height.  If a nonzero value is specified
 +
                      a linear falloff in height to zero at the edges of the filter is
 +
                      assumed.
 +
  -m: when -m is specified the filter's intensities must fit under the
 +
image intensities without any intensities going negative. If this
 +
not the case, then a zero value is put in that center pixel location.
 +
For example, if we specify a filter which is 100 brighter in the
 +
center than at the edge, and the filter is over a region that is 50
 +
brighter in the center, then (if -m is not specified) that pixel value
 +
might stay 50 (since the template can fit under the image intensity by
 +
lowering itself so its edges have negative values).  If -m
 +
is specified, the filter will not fit and the pixel will get set to 0.
 +
  -f #:  morphological filter shape. 6 = jack (ie, 3D cross, default)
 +
7= 3D asterisk, 4 = 2D plus
 +
1= 2D filled square, 8 = 3D cube, 2= 2D filled circle,
 +
3= 2D asterisk, 5= 2D cross.
 +
2D shapes are done one zslice at a time.
 +
  -l #: just use a single LINE, of specified orientation .
 +
1 = LL-UR, 2 = UL-LR, 3 = Left-Right, 4 = Up-Down.
 +
5 = In-Out in z, 6 = ULIn-LROut (a diagonal in 3D).
 +
  -s # # #:  size of x,y, and z "radius"  in pixels (4 4 2 default).
 +
  [-e|-d|-o|-c]:      erosion (min), dilation (max), opening (e then d), or
 +
closing (d then e). default is closing.
  
−
options:
+
note: entire image name is NOT required.
−
  -dx=# x coord at fusion time can differ by this and still be considered same path (default = 0)
+
note: a - in place of an image name means stdin or stdout.
−
  -dy=# y coord at fusion time can differ by this and still be considered same path (default = 0)
+
note: the origin of the filter element is at its center.
−
  -df=# fusion time can differ by this and still be considered same path (default = 0)
 
  
−
  -debug
+
see also: morph  - this is an older, one z-slice at a time version.
−
  -Debug
 
 
   
 
   
−
===  trackinfo.pl ===
+
</nowiki>
 +
===  multi_gauss_deriv ===
 +
<nowiki>
 +
 
 +
This program takes an image, blurs it with a Gaussian of
 +
specified standard deviation and then takes derivatives of the image
 +
It does all of this in the frequency domain.  The FFT will be done in parallel
 +
unless the environment variable NPROCS is set to 1.
 +
FFT_MAXMEM variable defines the maximum memory the FFT will use.
 +
outfile is the root name for images and/or pts (-m or -M) files.
  
−
# analyze output from "track or track -r".
+
Usage: multi_gauss_deriv [options] infile.i2i outfile
−
# assumes coords passed are zero indexed.  
+
options:
 +
  -x order:  number of derivatives in the x direction.
 +
default = 0
 +
  -y order:  number of derivatives in the y direction.
 +
default = 0
 +
  -z order:  number of derivatives in the z direction.
 +
default = 0
 +
  -s sigx sigy sigz:  standard deviation in spatial domain of Gaussian.
 +
  -S sigx sigy sigz:  standard deviation in frequency domain of Gaussian.
 +
default = 3.402823E+38  3.402823E+38  3.402823E+38
 +
  -l # res2 res3 ... :  # = number of different resolution levels to calculate.
 +
res2 ... res# are the relative sigmas for each resolution level after
 +
the first one. eg, -l 3 .9 .7  means that 3 blurred levels should
 +
be calculated and the sigma (in the freq domain) of the second
 +
level should be .9 of the first level (ie, more blurred), and the 3rd
 +
level should be .7 of the first level (as specified with -S).
 +
  -L # factor:  # = number of resolution levels. factor is the relative sigmas in
 +
the spatial domain for each level relative to the previous level.
 +
  -e epsilon:  cutoff for Gaussian intensity calculation.
 +
default = 0.040000
 +
  -r: apply scalefactor to output so min= -32766 or max= 32766. This is useful
 +
especially when all the values would otherwise be < 1.
 +
This also prints to stderr the min and max values before rescaling.
 +
  -R #: rescale output by multiplying all values by #.
 +
  -m: calculate coords of 3D maxima after blurring.
 +
  -M: same as -m, but also prevents writing out blurred images.
 +
  -n #: maxima must have an intensity > # (must use -m or -M with this)
 +
  -f #: max pixel must be > all neighbors by at least # to be a maxima. default = 0.000100
 +
note: image dimensions which factor to 2, 3, or 5 go fastest with the FFT.
 +
note: the entire image name is NOT required.
 +
note: compressed images will be uncompressed.
 +
note: a - in place of an image name means stdin or stdout.
 +
see also: max3d, gauss_deriv_unc, blur_spatial, canny, zerosu.
 +
 +
</nowiki>
 +
===  objs_via_ascent  ===
 +
<nowiki>
  
−
# usage:  trackinfo.pl -x=# -y=# [options] file.rpts
+
Associates each voxel in inimage.i2i which is > threshold with a 3d maxima in the image.  It finds
 +
the 3d maxima by doing a steepest ascent walk in the image.  Writes info about regions found to
 +
stdout in the same format as countobjs.  Writes an image with each pixel labeled with its region
 +
id to outimage.i2i.  
  
−
#  options:
+
Usage:
 +
  objs_via_ascent [options] inimage.i2i outimage.i2i > inimage.count
  
−
#    -reg=# : just print info about the specified region number
+
Options:
−
#    -track=# : just print info about the specified track
+
  -t threshold:any voxel > threshold in inimage.i2i is tracked to a 3d maxima (default = 0.000000).
−
#    -trackcoords=minsize:maxsize:miniod:maxiod:minlife:maxlife
+
  -i miniod maxiod : erase all objects whose iod doesn't fall in the specified range
−
# : print the coords (rpts format) of all tracks which satisfy
+
  -s minsize maxsize : erase all objects whose size (in voxels, inclusive) doesn't fall in the specified range
−
#   the criteria (or all its regions satisfy them). lifetime is # of timepts it exists for.
+
 
−
#    -tdist=mindist:maxdist: use with -trackcoords to specify min and max (xy manhattan) distance track travels
+
Examples:
−
#    -micro:   print coords of all