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Concept
Immunohistochemistry
Summary
Immunohistochemistry (IHC) is the most common application of immunostaining. It involves the process of selectively identifying antigens (proteins) in cells of a tissue section by exploiting the principle of antibodies binding specifically to antigens in biological tissues. IHC takes its name from the roots "immuno", in reference to antibodies used in the procedure, and "histo", meaning tissue (compare to immunocytochemistry). Albert Coons conceptualized and first implemented the procedure in 1941.
Visualising an antibody-antigen interaction can be accomplished in a number of ways, mainly either of the following:
Chromogenic immunohistochemistry (CIH), wherein an antibody is conjugated to an enzyme, such as peroxidase (the combination being termed immunoperoxidase), that can catalyse a colour-producing reaction.
Immunofluorescence, where the antibody is tagged to a fluorophore, such as fluorescein or rhodamine.
Immunohistochemical staining is widely used in the diagnosis of abnormal cells such as those found in cancerous tumors. Specific molecular markers are characteristic of particular cellular events such as proliferation or cell death (apoptosis).
Immunohistochemistry is also widely used in basic research to understand the distribution and localization of biomarkers and differentially expressed proteins in different parts of a biological tissue.
Preparation of the sample is critical to maintaining cell morphology, tissue architecture and the antigenicity of target epitopes. This requires proper tissue collection, fixation and sectioning. A solution of formalin is often used to fix tissue, but other methods may be used.
The tissue may then be sliced or used whole, dependent upon the purpose of the experiment or the tissue itself. Before sectioning, the tissue sample may be embedded in a medium, like paraffin wax or cryomedia. Sections can be sliced on a variety of instruments, most commonly a microtome, cryostat, or vibratome. Specimens are typically sliced at a range of 3 μm-5 μm.
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