Cell shape identification

Hello Everybody,

         I am studying the cell shape during morphogenesis of epethelial thissues. I have used GFP to visualize the cell-cell contacts by labeling E-Cadherin. The problem which I am facing is that I am not able to get cell shapes using any of the image software (including CellProfiler). In the picture, taken from microscope the cell-cell contacts are quit visible in green. Most of them are polygons and some of them are not. I want the skeletonized image of the pcitures I took from microscope. Any help in this context is appreciated. If you want I can give the pictures of my cells also.

with best regards
Vishu

Hello Vishu,

Can you please attach an image and your CellProfiler pipeline so far?

Best,
David

Hello David,

The image and the cellprofiler pipeline is attached below.
mdck.cp (7.05 KB)

Hi Vishu,

I attached a new pipeline. These are the basic changes:

  • I replaced EnhanceEdges with EnhanceOrSuppressFeatures. Not for any theorteical reason, but FindEdges is often hard to “tune” and EnhanceOrSuppressFeatures can often enhance edges, even though it wasn’t quite designed for that. I used the “Speckles” Feature Type here, though I expected “Neurites” to work better, but again, empirically, it didn’t.
  • Then ID the cell edges with IDPrimObjects. I changed the thresholding method to RobustBackground because Otsu was apparently being thrown off by a lot of pixels near zero and setting my threhold far too low. Otsu was assuming that this zero peak was the background, but it was just the low end of the background. RobustBackground will throw out these low intensity pixels from the threshold calculation
    ** I also removed any “clumped objects” processing. You want to keep the objects (cell membranes) intact here.
    ** I lowered the threshold correction factor, but if you need the membranes to connect, try lowering this further.
  • ConvertObjectsToImages: Necessary since Morph needs an image as input (binary here)
  • Morph: Skeletonizes the binary image, then de-spurs to get rid of “spur” artifacts

I didn’t look at the pipeline below that, so be sure to inspect it.

Let us know if that helps.
David
PIPE.cp (7.44 KB)

Hello David,

Thank you very much for your kind help. The pipeline you posted is doing a good job.

best regards
Vishu