Radioimmunoassay of colchicine with antisera exhibiting variable cross-reactivity

Colchicine-specific antibodies were produced in either goats or rabbits immunized with three different colchicine haptens conjugated to bovine serum albumin (BSA) at different coupling sites on the three rings of colchicine. Antibodies exhibited a variable cross-reactivity for metabolites and structural analogs of colchicine, which were dependent on the site at which colchicine coupled to the protein carrier. Specificity was also checked on urine samples by separating metabolites using high-performance liquid chromatography (HPLC) and radioimmunoassay (RIA) in tandem. The three antisera presented similar high-affinity constants for colchicine of the order of 10(10) M-1. A sensitive RIA for plasma colchicine was developed with each antiserum. The limit of detection of the three RIAs was 0.2 ng/ml. The inter- and intraassay coefficients were < 13%. RIA was linear up to 8 ng/ml. This RIA procedure was used to study the pharmacokinetics of a single dose of 1 mg oral colchicine in healthy volunteers and the colchicine concentrations of 27 plasma samples from patients on long-term colchicine treatment. No significant differences in plasma colchicine concentrations using the three assays were observed. This RIA procedure appears suitable for plasma colchicine pharmacokinetics and monitoring investigations.