A specific ELISA method for determining chloroquine in urine or dried blood spots

Abstract

Reported is an enzyme-linked immunosorbent assay (ELISA) that has been optimized and validated for the determination of chloroquine in urine or dried blood spots. The assay employs antisera raised in sheep to a chloroquine derivative conjugated to keyhole limpet haemocyanin and chloroquine conjugated to porcine thyroglobulin adsorbed onto the wells of a microtitration plate. The competitive binding of the antiserum to the wells was monitored using an alkaline-phosphatase-conjugated second antibody and a specific substrate. The assay exhibits no cross-reactivity with known chloroquine metabolites, other antimalarials, and commonly used drugs. The method was used to determine chloroquine in dried blood spot extracts and urine from a patient who was receiving a prescribed prophylactic chloroquine regimen. The drug was detected in the urine for 17 weeks and in the dried blood spots for 4 weeks after termination of the therapy.