Immunocytochemistry

Immunocytochemistry or ICC is a common lab practice which uses antibodies that target specific peptides or proteins antigens in the cell via specific epitopes. These bound antibodies can then be detected using one of several methods. ICC allows researchers to evaluate whether or not cells in a particular sample express the antigen in question. In cases where an immunopositive signal is found, ICC also allows researchers to determine which sub-cellular compartments are expressing the antigen.

Immuno"cyto"chemistry vs. immuno"histo"chemistry

Immunocytochemistry differs from immunohistochemistry in that the former is performed on samples of intact cells that have had most, if not all, their surrounding extracellular matrix removed. This includes cells grown within a culture, deposited from suspension, or taken from a smear. In contrast, immunohistochemical samples are sections of tissue, where each cell is surrounded by tissue architecture and other cells normally found in the intact tissue.

Methods

There are many methods to obtain immunological detection on tissues, including those tied directly to primary antibodies or antisera. A direct method involves the use of a detectable tag (e.g., fluorescent molecule, gold particles, etc., ) directly to the antibody that is then allowed to bind to the antigen (e.g., protein) in a cell.

Alternatively, there are many indirect methods. In one such method, the antigen is bound by a primary antibody which is then amplified by use of a secondary antibody which binds to the primary antibody. Next, a tertiary reagent containing an enzymatic moiety is applied and binds to the secondary antibody. When the quarternary reagent, or substrate, is applied, the enzymatic end of the tertiary reagent converts the substrate into a pigment reaction product, which produces a color (many colors are possible; brown, black, red, etc.,) in the same location that the original primary antibody recognized that antigen of interest.

Some examples of substrates used (also known as chromogens) are AEC (3-Amino-9-EthylCarbazole), or DAB (3,3'-Diaminobenzidine). Use of one of these reagents after exposure to the necessary enzyme (e.g., horseradish peroxidase conjugated to an antibody reagent) produces a positive immunoreaction product. Immunocytochemical visualization of specific antigens of interest can be used when a less specific stain like H&E (Hematoxylin and Eosin) cannot be used for a diagnosis to be made or to provide additional predictive information regarding treatment (in some cancers, for example).

External links

* [http://www.prosci-inc.com/Immunocytochemistry-Protocol.html Immunocytochemistry Staining Protocol]
* [http://www.cshprotocols.org/cgi/content/full/2008/2/pdb.prot4820 Immunohistochemistry of Whole-Mount Mouse Embryos]