Enzyme-Linked Immunosorbent Assay for Okadaic Acid: Investigation of Analytical Conditions and Sample Matrix on Assay Performance

Okadaic acid (OA) is a lipophilic marine biotoxin. In this study, OA was coupled with the carrier proteins keyhole limpet hemocyanin and bovine serum albumin as immunity and detection antigens by an active ester method. The polyclonal antibody against OA was prepared successfully, an indirect competitive ELISA (ciELISA) developed for the detection of OA in shellfish, and the reactive conditions of ciELISA optimized. The LOD (15% inhibition concentration) for the microwell plates was 1.28 +/- 0.38 ng/mL, corresponding to 12.8 +/- 3.8 ng/g. Two extraction methods were used to remove shellfish matrix interference with high recovery of spiked samples, and the methanol extraction of shellfish mussel was analyzed after dilution in phosphate-buffered saline. For validation of the optimized ciELISA, spiked and natural samples were analyzed by ciELISA, and HPLC with fluorescence detection. The correlation of linear regression equation was y= 1.0064x-10.234, and the correlation coefficient was 0.9347. From the results of the comparative study, the established ciELISA assay using polyclonal antibody against OA could be used in preliminary screening of suspicious shellfish samples.