|From:||"Coskran, Timothy M" |
Has anyone ever encountered the following type of IHC background problem:
With two separate antibodies, one a cytoplasmic marker and the other a
membrane marker, every single nucleus in the section turns brown. The stain
itself works, as most of the time you can read through the background, but
all test slides and negative controls (omission of primary) are affected.
Both antibodies are rabbit polyclonals, both are steam retrieved one with
citrate pH 6 the other with EDTA. Both are detected using a whole IgG
biotinylated goat anti rabbit secondary followed by Elite ABC and Dako
Liquid DAB+. Blocks included 3.0% hydrogen peroxide (aq), Dako Biotin
Blocking system and Dako Protein block. All were stained on an IHC stainer.
In addition, both of theses antibodies have been reliable for months (using
the above protocol steps), until now......
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