Running MeasureColocalization in CellProfiler (4.0.4).
Due to background noise I have binarized my images to only select for structure of interest. If I put this binzarized image as a maximum projection, the CellProfiler pipeline works. However, if I put the Zstack of the binarized image, the pipeline returns the error:
“Error while processing MeasureColocalization: local variable 'Thr_fi_c” referenced before assignment"
I thought it was an issue with the Zstack, but if I analyze the original grayscale (non-binarized) images, the pipeline works. I desire the Zstack because a maximum project results in seeming co-localization but there is clear separation in Z between the objects.
Has anyone encountered a similar issue or know why this occurs.