Streptavidin has a high affinity for biotin, with Kd = 1 x 10E-15 moles/L (Green, N.M. [1975] Adv. Protein Chem. 291:85). Streptavidin covalently conjugated to a reporter such as a fluorochrome (R-PE or FITC) or an enzyme (HRP or alkaline phosphatase) has utility in detecting, localizing, and quantitating biotin. Bound streptavidin conjugates can be detected either by fluorescence microscopy or by adding color producing enzyme substrate. This technique is especially useful with biotinylated antibodies. The biotinylation of antibodies is, relatively speaking, an easy procedure and the resulting product will continue to bind to its target antigen with a high degree of specificity and avidity. However, biotin itself cannot be readily detected. Biotin becomes highly apparent once streptavidin conjugated to a reporter is added.
Each streptavidin molecule can bind up to 4 biotin molecules. This feature is potentially useful for cross-linking biotinylated molecules.
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