I want to count number of multinucleated cells marked for nucleus (blue, Hoechst) and for cytosol (green, calcein). My cells are myotubes resulting from fusion of single nucleus myoblasts. Although it is easy to identify myotubes with nuclei forming clusters or chains (nuclei are overlapping or connected), some fused cells have separated nuclei. My intend is to identify cytosolic outlines (calcein) containing at least 3 nuclei.
My problem is that the cells are highly overlapped and I´m having problems to segment each cell outline. I upload an example of the cytosolic images and the nuclear mask. In my opinion, many fields (as that shown in the linked image) are simply too crowded to be segmented with fiji…
I would appreciate any help/opinion/experiences for this task.
Nuclear mask (previous to watershed):