Study of cytoskeletal changes induced by okadaic acid in HL-7702 liver cells and development of a fluorimetric microplate assay for detecting diarrhetic shellfish poisoning.

Diarrhetic shellfish poisoning (DSP) is a gastrointestinal illness with symptoms such as diarrhea, nausea, vomiting, headache, chills and moderate to severe abdominal pain. DSP has been recognized as a worldwide public health problem, causing great concern to the shellfish industry. Accumulation of DSP in shellfish is an unpredictable phenomenon that necessitates the implementation of a widespread collection and thorough monitoring program for mollusk toxicity. Therefore, development of accurate analytical protocols for the rapid determination of toxicity levels would be necessary. In this study we investigated cytoskeletal changes induced by okadaic acid in HL-7702 Liver Cells and developed a new cytotoxicity assay for detection and quantitation of DSP. This assay is based on fluorometric of F-actin depolymerization induced by okadaic acid (OA) compounds in HL-7702 liver cell line. The measurable range of OA was 2.5 ∼ 40 nmol/L. The detection limit of the F-actin assay for OA was 2.01 μg/100 g muscles in shellfish extracts. The performance of this assay has been evaluated by comparative analysis of shellfish samples by the fluorescent assay, mouse bioassay, and ELISA assay. Comparison of the results by all three methods revealed excellent consistency, the results of fluorescent assay were in significant correlation with ELISA assay (R(2) = 0.830). Examination of F-actin assay is very convenient, rapid, and sensitive, which can be used to quantify the amount of OA in shellfish samples.