A radioimmunoassay for determination of glibenclamide an other sulfonylureas

An antiserum was prepared for the determination of glibenclamide and for the estimation of other commercially available sulfonylureas. Rabbits were immunized with a glibenclamide-BSA conjugate. Tritiated glibenclamide was used as the tracer. The assay was performed in the presence of 8-anilinonaphthalenesulfonic acid to displace glibenclamide bound to serum protein, and free and antibody bound tracer were separated by dextran-coated charcoal. For glibenclamide determination in serum and plasma the limit of detection was 3 ng/ml. Sensitivity calculated for the whole determination range was 102 cpm for a 10 % concentration difference. Specificity studies showed a cross-reaction of less than 0.1 % for glibenclamide metabolite M1 and 9 % for metabolite M2. Other sulfonylurea drugs display cross-reactivities from 0.1% (chlorpropamide) to 190% (gliquidone). Both intra-assay and inter-assay imprecision were below 10 %. Accuracy was established by comparison of the present method with HPLC. The assay was applied to the specific determination of glibenclamide in clinical trials and for diagnosing factitious hypoglycemia caused by sulfonylureas.