Phase contrast image segmentation for counting fibroblasts

Posted by sfreema1 on Jul 28, 2020; 12:47pm
URL: http://imagej.273.s1.nabble.com/Phase-contrast-image-segmentation-for-counting-fibroblasts-tp5023774.html

Good morning,

I am trying to find a workflow for counting cells in a phase contrast image.
These are fibroblasts that I grew in a 96-well plate. Several images were
stitched together.

As I look through other similar questions here and elsewhere, I find a lot
of the examples are geared towards more circular cells. I have tried using
the common workflow (thresholding/binarizing -> analyze particles) but I
have not been content with my results.

Because the images were taken in the 96-well plate, all my phase contrast
images have a feature where the centers of the wells are darker and have
more contrast, while as I move away from center, the features brighten and
lose contrast.

I found that Sharpen->Find Edges was helpful in outlining the cells and
apparently removing the uneven illumination, but then I have not been able
to proceed from there.

I feel another issue with these cells is that they crowd very closely
together, and I think it is hard to distinguish two separate cells even by
eye sometimes.

I would appreciate insight anyone has to give. I have attached a sample
image below from my dataset. A12_-1_1_1_Stitched[Phase_Contrast]_001.tif
<http://imagej.1557.x6.nabble.com/file/t382700/A12_-1_1_1_Stitched%5BPhase_Contrast%5D_001.tif>  



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