Enzymatic solid-phase assay for biotin and a biotin-benzodiazepine conjugate

A novel enzymatic ligand binding assay for biotin and its benzodiazepine conjugate is based on their binding to horseradish peroxidase-avidin conjugate (A-P) followed by the uptake of biotin-unsaturated A-P onto polystyrene beads coated with biotin-BSA. The detection limit is 1.3 x 10(-16) mol per tube (300 microL) with a 3.3 x 10(-12) M A-P solution and varies with the conjugate concentration employed. The coefficient of variation for 10 repetitive assays of 10(-15) mol of biotin is 6.22%.