Can anyone explain to me why I got a large amount of myelin staining when I did IHC for Bax (Ser184) using the ABC method and DAB? I know Bax is associated with apoptotic neurons, which is what I was looking for (and did find a few nicely labeled cells), but why do I also have so much labeling of myelin?
I think your picture shows unspecific staining of blood vessels because of bad perfusion. try to inject more PBS to get rid from blood completely.
Good luck
Dear Vincent,
I agree with Imam in that it looks like blood vessels. You might be able to get rid of the endogenous reaction of DAB with red blood cells by incubation new sections in a higher concentration of H2O2 (0.5%) before immunolabeling, but I haven’t had much luck with it. I suggest that if you need to use any remaining sections from these animals, you try swithching to an amplification system that doesn’t rely on HRP. Are you able to use fluorescence? If so, you can replace the ABC with a fluorescently-tagged streptavidin.
Good Luck!
Jill
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