Zebrafish embryos were fixed overnight at 4C when they had reached 60% epiboly. They were processed for ICC/IF as outlined above and post-fixed in PFA for 20min at r.t. after extensive washing of the secondary antibody. Pieces of lateral blastoderm were then mounted in Vectashield and image stacks through the epi- and hypoblasts were collected on a confocal microscope. The images shown are maximum intensity Z-projections of 4, 1micron slices. The embryonic cell cycle has minimal gap phases at this point, so most nuclei would be expected to be PCNA-positive. The left panel shows DAPI stained nuclei, the center panel is PCNA staining, and the right panel is the merged image.
Mr. Hank Farr
Submitted Mar 30 2012
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