Anti-warfarin antibody preparation and its characterization for radioimmunoassay

Article abstract of DOI:10.1111/j.2042-7158.1982.tb04750.x

Anti-warfarin antiserum was prepared in rabbits by immunization with a synthesized warfarin antigen, 4'-azo-warfarin human serum albumin, which possesses two enantiomorphic haptenic sites of warfarin on the molecule. The antiserum recognized both R- and S-warfarin to the same degree, 50% cross reactivities of racemic warfarin, respectively. One of the warfarin metabolites, racemic warfarin alcohol, showed 1% cross reactivity, and R- or S-warfarin alcohol have half the reactivity of the racemic alcohol. Rabbit plasma warfarin levels were determined by radioimmunoassay using this antiserum and racemic [¹⁴C]warfarin and by fluorometric assay after isolation by thin layer chromatography. After a single administration of warfarin (2 mg kg^-1 orally or 400 μg kg^-1 i.v.), the plasma levels determined by both assay methods showed a good correlation (r = 0.97, P < 0.001, Y = 1.04-0.09). The results show that the radioimmunoassay can determine total plasma warfarin without interference of plasma metabolite. The applicability and limitation of the radioimmunoassay for pharmacokinetic study are discussed.