I’m new to CellProfiler and I have a set of tricky pictures to count. In a nutshell, we’re doing simple invasion in collagen-coated chambers and then staining with crystal violet. One problem is that our cell type has a tendency to clump and overlap along with having long processes that touch neighbors. I’ve been tinkering with multiple types of pipelines and am still having trouble getting the program to recognize the unusual cell shapes in a homogenous manner–additionally, I don’t know how to mask for the little air pockets in the collagen. I’ve included a picture below and some of the previous (simplistic) pipeline that has worked the best for me so far.
One problem I’ve also been having is that I would try to select for the red channel, apply a binary threshold and then count the resulting dark clumps but for some reason the program is finding random things in the white background to count as opposed to the stained cells. Is there a way to invert this so that it will count the dark and not the light?
- Crop (1-1000 from each edge, discard edges)
- Color to Gray (combine)
- Apply Threshold
- above threshold
- number of pixels to expand: 0.1
- Global, otsu, 2 classes, weighted variance, no smoothing
- Identify Primary Objects
Not sure where to go from here, but I feel like I’m probably missing a few important modules somewhere.
Thanks for reading!