Abstract
The identification of antigenic substances with antibodies can only occur through the use of a reporter molecule. One way of doing this is through the use of enzymes. Enzymes act upon a substrate and that substrate, or a molecule affected by that substrate, in turn becomes detectable by a variety of methods. There are many enzymes available for this purpose. The most common is peroxidase. Another widely used enzyme is alkaline phosphatase. Each enzyme has a few chromogenic substrate solutions with which it can react to change a color visualized through the use of selected instruments, including the microscope. Antibodies can be labeled with an enzyme directly, or secondary antibodies can be labeled with the enzyme and employed in an indirect technique. Also, immunoglobulin labeled polymers labeled with enzyme can be used and the enzymes themselves can serve as antigens in immunoenzyme complex procedures. Finally, avidin or biotin can be labeled with enzyme, and used either singly or in complexes, and peroxidase mediated biotin amplification can be used to increase the sensitivity in some procedures.
The opinions or assertions herein represent the personal views of the author and are not to be construed as official or as representing the views of the Department of the Army or the Department of Defense.
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References
Nakane P, Pierce G (1966) Enzyme-labeled antibodies: Preparation and application for the localization of antigens. J Histochem Cytochem 14:929–931
Farr A, Nakane P (1981) Immunohistochemistry with enzyme labeled antibodies: A brief review. J Immunol Meth 47:129–144
Swanson P (1988) Foundations of immunohistochemistry. Am J Clin Pathol 90:333–339
Ayala E, Martinez E, Enghardt M, Kim S, Murray R (1993) An improved cytomegalovirus immunostaining method. Lab Med 24:39–43
Sakanaka M, Magari S, Shibasaki T, Shinoda K, Kohno J (1988) A reliable method combining horseradish peroxidase histochemistry with immuno-β-Galactosidase staining. J Histochem Cytochem 36:1091–1096
Flugelman M, Jakitsch M, Newman K, Casscells S, Bratthauer G, Dichek D (1992) In vivo gene transfer into the arterial wall through a perforated balloon catheter. Circulation 85:1110–1117
Musiani M, Zerbini M, Plazzi M, Gentilomi G, LaPlaca M (1988) Immunocytochemical detection of antibodies to Epstein-Barr virus nuclear antigen by a streptavidin-biotin-complex assay. J Clin Microbiol 26: 1005–1008
Bos E, van der Doelan A, van Rooy N, Schuurs A (1981) 3,3’,5,5’-tetramethylbenzidine as an Ames test negative chromogen for horse-radish peroxidase in enzyme-immunoassay. J Immunoassay 2:187–204
Hsu S, Soban E (1982) Color modification of diaminobenzidine (DAB) precipitation by metallic ions and its application for double immunohistochemistry. J Histochem Cytochem 30:1079–1082
Sternberger L (1979) Immunocytochemistry. Wiley, New York, NY
Hinglais N, Kazatchkine M, Mandet C, Appay M, Bariety J (1989) Human liver Kupffer cells express CR1, CR3, and CR4 complement receptor antigens. Lab Invest 61:509–513
Tse J, Goldfarb S (1988) Immunohistochemical demonstration of estrophilin in mouse tissues using a biotinylated monoclonal antibody. J Histochem Cytochem 36:1527–1531
Hsu S, Raine L, Fanger H (1981) Use of Âavidin-biotin-peroxidase complex (ABC) in immunoperoxidase techniques: A comparison between ABC and unlabeled antibody (PAP) procedures. J Histochem Cytochem 29:577–580
Swanson P, Hagen K, Wick M (1987) Avidin-biotin-peroxidase-antiperoxidase (ABPAP) complex. Am J Clin Path 88:162–176
Bobrow MN, Litt GJ, Shaughnessy KJ, Mayer PC, Conlon J (1992) The use of catalyzed reporter deposition as a means of signal amplification in a variety of formats. J Immunol Meth 150:145–149
Merz H, Malisius R, Mannweiler S, Zhou R, Hartmann W, Orscheschek K, Moubayed P, Feller AC (1995) A maximized immunohistochemical method for the retrieval and enhancement of hidden antigens. Lab Invest 73:149–156
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Bratthauer, G.L. (2010). Overview of Antigen Detection Through Enzymatic Activity. In: Oliver, C., Jamur, M. (eds) Immunocytochemical Methods and Protocols. Methods in Molecular Biology, vol 588. Humana Press. https://doi.org/10.1007/978-1-59745-324-0_24
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DOI: https://doi.org/10.1007/978-1-59745-324-0_24
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