Cell counting with layered cells




I have a set of images that pretty consistently are undercounting with my existing pipeline. I think it is because the different cells vary in intensity, but not quite sure. The cell set, which i have attached, should come to 125 unique cells in the blue channel according to the hand counted value, but regardless of adjusting the typical cell size, the IDP/tracking is coming up with around 85 cells. The pipeline I have been using is attached. Any help or direction to look would be very much appreciated. output.cppipe (17.0 KB)
dapi_images.zip (6.5 MB)


One thing you can try adding to your pipeline is CorrectIlluminationCalculate and CorrectIlluminationApply to help reduce the issue of variation in intensity. In addition, resizing and filtering (e.g MedianFilter) the image prior to segmentation often helps. See attached draft where you can possibly start and see if that helps!
VCoutputVC.cppipe (19.7 KB)