PgFocus Calibration

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How to calibrate pgFocus

  1. PgFocus must be connected to a piezo controller, which is connected to a piezo positioner.
    • PgFocus assumes that 1 Volt moves the objective 10µm.
  2. Focus on a coverslip as you would normally.
    • Cells are not needed but the media should be same.
    • A focus position must be able to be detected by pgFocus, and it should be roughly centered on the Linear Light Array (LLA). Anywhere between 50-70 offset should be fine.
  3. In the Stand-alone pgFocus application, type the letter 'c' to activate the calibration process.
    • You can also use the pgFocus µManager plugin.
  4. PgFocus will move the objective until the focus position reaches 100 offset on the LLA.
    • This will determine the maximum voltage that pgFocus will use during the calibration process.
    • pgFocus will either use the maximum voltage as determined by this step, or voltage needed to move the objective 3000nm up, whichever is less.
  5. PgFocus will then slowly lower the objective by 500nm until it reaches -3000 nm or the bottom the linear array sensor (28 offset).
    • Voltage and LLA offset will be collected at each step.
  6. PgFocus will then perform a linear regression on the data points it collected, and save the result in the eprom.

500 nm is hard coded based on DAUPERVOLT/MICRONPERVOLT/2 = 81.9. It shouldn't be hardcoded... : D