  Previously, we have identified the cytoplasmic zinc metalloprotease
insulin-degrading enzyme(IDE) in human tissues by an immunohistochemical method
involving no antigen retrieval (AR) by pressure cooking to avoid artifacts by
endogenous biotin exposure and a detection kit based on the labeled
streptavidin biotin (LSAB) method. Thereby, we also employed 3% hydrogen
peroxide(H2O2) for the inhibition of endogenous peroxidase activity and
incubated the tissue sections with the biotinylated secondary antibody at room
temperature (RT). We now add the immunohistochemical details that had led us to
this optimized procedure as they also bear a more general relevance when
demonstrating intracellular tissue antigens. Our most important result is that
endogenous peroxidase inhibition by 0.3% H2O2 coincided with an apparently
positive IDE staining in an investigated breast cancer specimen whereas
combining a block by 3% H2O2 with an incubation of the biotinylated secondary
antibody at RT, yet not at 37 degrees Celsius, revealed this specimen as almost
entirely IDE-negative. Our present data caution against three different
immunohistochemical pitfalls that might cause falsely positive results and
artifacts when using an LSAB- and peroxidase-based detection method: pressure
cooking for AR, insufficient quenching of endogenous peroxidases and heating of
tissue sections while incubating with biotinylated secondary antibodies.
