When brightfield (right?) objects are clumped, they are very hard to declump. I tried changing some settings in IDPrimary (increasing the threshold correction factor, and raising the suppress local maxima), but it didn’t improve that much. You could tweak all you want for one image, but I don’t predict that it will be very robust. I also tried some Morphological filtering (“Morph” module in CP) using the Open operator , but the little intensity depression in the middle of your cells causes issues with this approach (as well as the current approach).
Some thoughts come to mind:
(1) Add a fluorescent marker, preferably for a single subcellular compartment (nucleus, etc). I understand if that is not your first choice, but it is the most straightforward one.
(2) Instead of using the Combined image in ColorToGray, try using the red channel only using the Split option. To my eye it seems (very) marginally cleaner than the combined image.
(3) You could try UntangleWorms. These modules were developed for C. Elegans, but they might work for your images too. There is some more work involved than your current pipeline (defining a “Worm” model, etc) but it might work if you are stuck with bright field.