THE SA RABC TECHNIQUE - A NEW ABC PROCEDURE FOR DETECTION OF ANTIGENSAT INCREASED SENSITIVITY

Since its introduction, the avidin-biotin-peroxidase (ABC) complex has become an invaluable detection system for a wide variety of bioanalyt ical applications. In these techniques, avidin and biotin-peroxidase a re mixed at a pre-determined ratio so that the soluble ABC complex ret ains biotin binding sites. Consequently, the complex contains an exces s of avidin over biotinylated peroxidase residues. On theoretical cons iderations, however, an ABC complex designed for maximal signal intens ity must consist of an excess of peroxidase over avidin molecules. The refore, ABC complexes with reversed molar ratios of biotinylated perco xidase to avidin (rABC complexes) were prepared and an intermediate st reptavidin step was introduced to bind the rABC complexes to biotinyla ted IgG molecules. The signal generating power of this new streptavidi n-rABC sequence proved superior to that of the conventional ABC techni que in ELISA assays and in immunostaining of tissue sections.