The Use of Antiavidin Antibody and Avidin-Biotin-Peroxidase Complex in Immunoperoxidase Technics

Avidin has an extraordinary affinity for the small-molecule vitamin biotin. Covalently coupling biotin or avidin to peroxidase molecules does not interfere with their normal biochemical functions. The avidin or biotin molecules, either peroxidase conjugated or unconjugated, can be brought to the antigen sites by means of an antiavidin antibody. Several immunohistochemical staining technics based on this principal have been described. The method utilizing an avidin-biotin-peroxidase complex was found to be more sensitive than the unlabeled antibody (PAP) method. This method involved four sequential staining procedures: (1) primary antibody (goat anti-human antigen); (2) secondary antibody (rabbit antigoat IgG) added in relative excess; (3) goat antiavidin antibody; (4) avidin-biotin-peroxidase complex. The applications of this technic are discussed.