Endogenous biotin

Handling HIER induced demasking of biotin

Using avidin-biotin based visualization systems, false positive staining frequently appears after heat induced epitope retrieval (HIER) due to the reactivity of endogenous biotin. As a general experience, the more efficient HIER, the more biotin is retrieved. Biotin is present in mitochondria of metabolically active cells. Particularly parenchymal cells of the liver, kidney, colon, thyroid and breast, as well as the corresponding carcinomas are prone to give this staining reaction.

Endogenous biotin blocking can be carried out using protocols based on either avidin (or egg white) and biotin (or skim milk). A good alternative is the use of a non-biotin based visualization system, e.g., EnVision+.