I have been trying to design a pipeline to count RPE cells and measure their area. Some problems we have been having are that they are multinucleated, there are nuclei on the cell borders, and that they have vastly different cells size and shape (on the knockouts). I originally tried to use the identifyprimary and then identifysecondary with propagation, but could not get an accurate number.
So the pipeline I am currently using works like this:
- Take the nuclear staining (blue) and use identifyprimary on the nuclei
- Make an image of the nuclei
- Delete nuclei from the inverted image of the cells+nuclei (green)
- IdentifyPrimary function looking for the cells (cytoplasm)
It works well for the wild type images, usually with about 3-5% error, but is very inaccurate with the knockouts. It often counts the large cells as multiple cells or has problems separating elongated cells.
I have attached some images of the wild type and KO, as well as my current pipeline. Any help or suggestions would be greatly appreciated.
RPE Pipeline.cppipe (12.1 KB)
Edit: Here is a link to the images, because they didn’t seem to upload correctly:
Google Drive Link