A direct method of radioiodination of biotin derivatives is presented whereby radiolabeled sodium iodide is exchanged with a tributylstannyl group under oxidative conditions. The in vitro binding of each radioiodinated biotin derivative to streptavidin is saturable and three of the compounds exhibit a strong affinity for streptavidin. All the biotin-streptavidin complexes are rapidly catabolized in human serum.