Hi CellProfiler team,
Thanks a lot for this open-sourcing program, which makes my imaging quantification possible.
I’m trying to process my imaging assay data acquired by automatic imaging plate reader by CellProfiler. As seen in my attached files, there’re two images stained by IF, one shown with protein enriched in plasma membrane and another with protein diffused in cytoplasm. And there’re two images with DAPI staining. I had a pipeline by which I shrunk cells for 4 pixels to identify plasma membrane and cytoplasm with the rest and then calculate the ratio between them. I feel segment of plasma membrane with this way is ok, however the results of ratio of PM to cytoplasm are only 1.14 and 1.02 respectively for those two image set. Then I tried to run your pipeline example with cytoplasm-nuclear location (ExampleVitra). For this example one image has staining in cytoplasma and another in nuclei, however the results as shown by ratio also gave a small difference. I also attach this example with image, pipeline and results. I did a little modification for this pipeline by adding illumCorrection in the same pipeline.
I’m confused with these results if I miss some points. I wonder if I should set a threshold to subtract background. Could you please review pipelines and have suggestions for me?
Thanks a lot.