Sample image and/or code
We are trying different IF markers of cell membranes to help with segmentation (otherwise impossible), and this one looks absolutely amazing! This is in the liver, where multi-nucleated cells mean that nuclei-based segmentation does not work.
Segment my cells
When I try to segment in cell profiler, I’ve not been able to find settings that satisfactorily segment my cells, even though the edges seem obvious (dark black lines in the image above). For example, about one third left from the right edge, one third down from the top edge are three cells that have randomly been divided into 2 or 3. Since most segmentation is done using nuclei, these are the steps I took which might be part of the problem.
- Max projection
- split channel to only my border marker
- Negative of image
Settings: Here are setting used for the IdentifyPrimaryObjects segmentation above. But, I have tried many combinations of threshold strategy, method, smoothing, correction factor, window sizes. It seems the problem has to do with the declumping, but I haven’t been able to improve the segmentation much better than the image above. I tried making a binary mask in ImageJ and the segmenting algo still draws lines through the middle of cells!
Thank you for considering!