[Histonet] Problems with c-fos immediate early gene labelling
I'm hoping I could get some helpful advice on what might cause my c-fos
IHC labelling to work well in one lab, but not my own. I am running the
protocol on rat brains using a hydrogen peroxide/methanol step for
endogenous peroxidase blocking, a normal goat serum blocking step, a
c-fos Ab-5 primary from oncogene, a goat-anti rabbit biotinylated
secondary, a streptavidin horeradish peroxidase, and a nickle enhanced
Unfortunately, although I have all of the exact same antibodies, chemicals,
and concentrations as the lab from which the protocol was developed, the
label is weak and not very specific when I run it at my home lab. In fact, I
even took brain slices from the same rats that I was running at my lab and
ran them at the lab where the protocol was developed, and found that the
c-fos label was beautiful with a low background.
Does anyone has any suggestions as to why this might be? One person
at the other lab suggested that possibly the wells that we run the tissue in
were not being cleaned properly (we soak them in bleach while the other
lab uses just pex). Could this have any effect?
Thanks for any advice you can provide!!
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