I am very new to using Cellprofiler.
We perform big genome-wide RNAi screen in Drosophila S2R+ cells in 384-wells plates. Cells were stained with DAPI (channel2 in my images names) and anti-tubulin-FITC antibodies (GFP channel, channel1 in my images names).
We want to analyse the images taken with 20x magnification.
The thing is that some of our cells at some experimental are very big cells, and some at other conditions are very small and crowded. But for this I was able to create a pipeline which allows me to find both, small and large cells, so it was not such a big problem.
The problem which I have is to distinguish the very small nuclei from the dirt and cell debris, which I have in wells in which cells totally died out (example: positive control wells, images channel1_2 and channel2_2). I tried to rule out them by playing with size limits and threshold, but I was unsuccessful so far, when it was getting better with excluding debris, I started also exclude the proper (but very small) nuclei from other images. Do you have any ideas what should I do?