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The Avidin–Biotin Complex (ABC) Method and Other Avidin–Biotin Binding Methods

Part of the Methods in Molecular Biology book series (MIMB,volume 588)


Immunoenzyme methods can be enhanced by the use of the high affinity molecules, avidin and biotin. The binding of avidin to biotin is almost irreversible. By labeling a detection enzyme such as horseradish peroxidase with biotin, and a secondary antibody (reactive against the antigen detecting primary antibody) with biotin as well, these two compounds can then be linked irreversibly with avidin. For this process, the biotinylated enzyme is complexed with avidin in solution and this avidin–biotin complex (ABC) is then introduced to the biotinylated secondary antibody, where it binds to primary antibody-antigen sites. Also, enzyme-labeled avidin molecules can be used to bind biotinylated secondary antibodies with greater resolution. Finally, biotinylated tyramide used in conjunction with peroxidase precipitates even greater amounts of biotin molecules for detection by enzyme-labeled avidin molecules.

Key words

  • Avidin
  • Biotin
  • Avidin–biotin complex (ABC)
  • Labeled avidin binding (LAB)
  • Biotinylated tyramide
  • TBS buffer
  • DAB chromogen

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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  • DOI: 10.1007/978-1-59745-324-0_26
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Correspondence to Gary L. Bratthauer .

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Bratthauer, G.L. (2010). The Avidin–Biotin Complex (ABC) Method and Other Avidin–Biotin Binding Methods. In: Oliver, C., Jamur, M. (eds) Immunocytochemical Methods and Protocols. Methods in Molecular Biology, vol 588. Humana Press.

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