I need to do spot detection and need to compare the ratio of the number of spots detected in one image versus number of spots detected in another image. For that reason, I want to keep the background subtraction equal for spot detection in both images, but Imaris (9.3.0) does not give any info on how this background is calculated and therefore is probably different across images and as such will influence the number of detected spots.
Does anybody know how this background subtraction works in spot detection?
Use ImageJ that is a GUI so you can manipulate and use various operators !!
Subtraction shows the difference in the frames , if the spot wasn't there then that can be seen (it might require more at times)
I usually use ImageJ to do manual background subtraction, but maybe you can do this in Imaris. Draw an ROI in an area of background and measure the mean intensity in this ROI. Then subtract this value from all pixels in the images (Process, Math, Subtract). Worse case, you could do this subtraction step in ImageJ and then proceed to Imaris.
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