So I have to analyze punctuate fluorescence (see example image in red) in cells that expressed EGFP fluorescence (see example EGFP image).
My images are from a Z-stack obtained by confocal microscopy. In my initial image, I split the green and red channels, and I do a Z-project for each channel (Max intensity) followed by substraction of background in red and green channels.
Then, I threshold the red channel followed by “analyze particles” tool, and it works pretty well.
The problem I have is that I only want to analyze particles that are in a given overlay (EGFP cell contour I draw in the green channel). I didn’t find how to restrict analysis of red fluorescence in all of my contoured EGFP cells.
For the moment, I contour individually each EGFP cell, then Ctrl-MajE in the red channel, and finally analyze the particles in the given contour using the ROI I created with the thresholded image. As you can imagin, this manipulation cell by cell is very time consuming and I wanted to automatize the analysis to measure the particles in all my contoured EGFP cells at once.
Sorry if my english is not good, I hope I has been understandable.
Thanks a lot,