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The lab utilizes the ABC method on all of its IHC staining protocols. Staining results are significantly improved by the pretreatment of the sections in an antigen retrieval soloution (DakoCytomation Target Retrieval Solution) under controlled heat and pressure using a Decloaking Chamber (Biocare Medical). The Target Retrieval Solution acts to break protein cross links formed by fixation in 10% Neutral Buffered Formalin, thus uncovering hidden antigenic sites.
Background staining is reduced by the application of a protein block (DakoCytomation Serum-Free Protein Block) directly to the sections to prevent non-specific background staining typically seen when performing IHC on animal tissue. Endogenous peroxidase activity, which is found in many tissues, is also blocked by the pretreatment of the sections with hydrogen peroxide prior to the application of the primary antibody.
The following is the immunohistochemistry protocol utilized by the Histology / IHC lab: