Nuclei Watershed Separation

Revision as of 04:15, 15 July 2009 by White (talk | contribs) (create tutorial for watershed segmentation of DAPI stained nuclei.)
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A very common biological sample for microscopy is DAPI stained DNA in cell nuclei. The staining delineates the nuclei pretty well, since in a metaphase cell there is DNA all over the nucleus. however, the staining is not homogenous, as there are areas of more or less condensation of the chromosomes. This makes the nuclei appear granular. Worse still, in may tissues that are interesting for developmental biology, the cells are tightly packed, and are composed mostly of nucleus with very little cytoplasm separating them. The nulclei often seem to touch each other, which in reality, of course, they can not. There is a membrane or two at least between them, but an optical microscope cant resolve that. The PSF is much bigger then the width of a membrane!

So, we are faced with the probelm of being able to separate apparently touching objects :-(

Luckily, there is a method for doing exactly that. The Watershed method.

Ok, so how can we segment, watershed (separate touching objects) and then count / measure the objects in FIJI? Read on...

  • open the sample image of DAPI stained nuclei