I have been running JACoP 2 on your screen capture, and here is the output:
Image A: Nuclei
Image B: Cells
Colocalization based on centres of mass-particles coincidence
Threshold for Image A=128; Image B=128
Particles size between 0 & 2391688
Image A: 10 centre(s) colocalizing out of 80
Image B: 8 centre(s) colocalizing out of 17
To do that, I’ve been using the “Centre-particles coincidence” option. The rational is that you have big objects on one channel and small one in the other one: you may want to summarize the small ones as points and see if they match with any of the underlying objects.
JACoP will compute where the centres are on image 1, and where the objects are on the image 2. It will then count the numbers of centres from image 1 falling onto objects from image 2.
From the exported data, you may see that you have 17 cells and 80 spots of nuclear marker.
When looking at the nuclei centres falling onto cells, you find 10 of them (out for 80) on cells and 8 (out of 17) centres of cells falling onto nuclear markers.
I hope this helps.
I’ve enclosed the latest version of JACoP (75.8 KB): it seems the Tudor website where it has long been hosted is down. I’ll put it on GitHub ASAP (I need to do a bit of cleanup on the user interface before).