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| - Development of a Sensitive Enzyme-Linked ImmunosorbentAssay for the Determination of Domoic Acid in Shellfish
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| - Polyclonal antibodies for domoic acid were generated from rabbits after the animals had beenimmunized with either domoic acid−keyhole limpet hemocyanin (KLH) or domoic acid−bovine serumalbumin (BSA). A competitive direct enzyme-linked immunosorbent assay (cdELISA) and a competitiveindirect ELISA (ciELISA) were used for the characterization of the antibodies and for analysis ofdomoic acid in blue mussels and clams. The antibody titers in the serum of rabbits immunized withdomoic acid−KLH were considerably higher than those in rabbits immunized with domoic acid−BSA. The antibodies from the rabbits immunized with domoic acid−KLH were further characterized.In the cdELISA, the concentrations causing 50% inhibition (IC50) of binding of domoic acid−horseradishperoxidase to the antibodies by domoic acid and a domoic acid analogue, kainic acid, were found tobe 0.75 and 200 ng/mL, respectively. In the presence of blue mussel matrix, the detection limit ofdomoic acid was <25 ng/g. The overall analytical recovery of domoic acid (25−500 ng/g) added tothe blue mussels and then extracted with 50% aqueous methanol in the cdELISA was found to be81.1%. The efficacy of cdELISA was also confirmed by the high-performance liquid chromatographymethod. Analysis of domoic acid in shellfish samples showed that 10 of the 15 shellfish examinedwere contaminated with domoic acid at levels of <50 ng/g. Keywords: Domoic acid; antibodies; ELISA; shellfish
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