Hi,

Thanks so much for the support so far! I’m trying to build a pipeline for quantification of co-localization and was hoping to clarify a few things about the modules. Basically, what I’m trying to measure at this point is the percentage of TUNEL-positive nuclei in a stack of images. My basic approach is to measure the number of nuclei objects and the number of TUNEL objects (nuclei blue and Tunel yellow-green) per section, then to use one of the colocalization modules to determine which TUNEL stains co-localize with nuclei. I’ve actually tried to use both the measurecorrelation and relate modules, although I have to be honest that I’m not exactly sure how to interpret the readout. I finally settled on ‘relate’ in the attached pipeline because I’m not as interested in the correlation of intensity, but rather of objects themselves, and this one I think gives me a column that tells how many Tunel per nuclei per section. I’m not entirely sure what to make of the correlation number (which ranges from 1E2 to -1E3), and how I would use this in my analysis. All I really want to know is what percentage of Tunel stain is co-localized with nuclei so that I can adjust my total counts per section to reflect ‘true’ Tunel-positive nuclei. For example, a section might have 5 Tunel objects and 150 nuclei, with .80 co-localization of Tunel with nuclei such that I could determine 4 Tunel-positive nuclei in that section. Unfortunately, I’m not exactly sure how to arrive at the 80% value based on either the measurecorrelation or relate modules, so if you have any advice, I’d greatly appreciate it.

One other question concerning stacks, do you have any suggestions for loading .avi files, and is there any way to determine that an object counted in 1 section of a stack will not be counted in others. I think this was addressed in another forum, although I haven’t been able to verify that the software is doing this in my analysis.

Thanks again for all your help!

Mike

TUNELPIPE.mat (1.29 KB)