Function: Light Program

Function: Light Program

The Light Program function allows creation of a custom sequence of laser illumination for imaging:
  1. Total Number of Steps: add a different sequence of laser states to the light program. The maximum number of steps allowed is 5.
  2. Number of States: add a different order of laser states to a step. The maximum number of states allowed is 25 per step.
  3. Repeat Continuously or Number of Times To Run: choose the repetition mode for each step.
  4. Groups: generate a virtual partition to separate localizations that occur in the same channel. This allows applying localization filters to fluorophores that overlap in spectra but are separate in frames.
  5. Save Frame: choose to whether save the frame or not to the local drive.
  6. Load From File/Save To File: load/save a light program from/to a JSON file.
  7. Clear: clear up the program.
  8. Save And Activate Program: save the current program in NimOS and activate it for acquisition.
  9. Save And Close: save the current program in NimOS and close the program window.
  10. Revert Changes And Close: revert the changes made to the current program and close the program window.

Instructions

  1. Click on Program to open the program window.
  2. In the pop-up window, set the desired laser steps and states.
  3. Click on Save And Close to save the program in NimOS.
  4. Set No. Of Images to the total number of frames specified in the program based on this equation:
  5. Check Enable Light Program and click on Acquire to start the acquisition.

Discussions

  1. For more detailed instructions on how to use the light program and how to interpret the result, check here.
  2. The effective laser power is not the same with and without the light program since lasers are switched on/off for each frame with the light program whereas the lasers are always on without the light program. Normally the laser power specified in the light program will be 10~30% higher than usual.
  3. All frames are acquired at the same exposure specified in Exposure (ms). An exposure of 20 ms is the recommended minimum, which corresponds to the hardware limit of the rolling shutter camera.