Radioimmunoassay of the anticonvulsant agent clonazepam

A simple and specific radioimmunoassay was developed for the determination of the anticonvulsant agent clonazepam directly in plasma without extraction. Antibodies to clonazepam were produced in rabbits after immunization with an immunogen prepared by covalently linking the 3-hemisuccinyloxy derivative of clonazepam to bovine serum albumin. When employing 3H-clonazepam as the tracer, the radioimmunoassay has a limit of sensitivity of 5 ng/ml using a 0.1-ml sample of plasma. The antibodies exhibited a high degree of specificity for clonazepam; no cross-reactivity was observed with its 7-amino and 7-acetylamino metabolites nor with a number of other widely prescribed anticonvulsant agents that might be administered in conjuction with clonazepam. Satisfactory agreement was obtained for the plasma levels of clonazepam in humans when samples were assayed by the radioimmunoassay and an established electron-capture GC technique. By virtue ot its simplicity, the radioimmunoassay offers a distinct advantage to the clinician for monitoring plasma clonazepam levels and the compliance of patients undergoing anticonvulsant therapy with the drug.