Development and Validation of an Immunochromatography Test System for Determining Okadaic Acid in Seafood

Okadaic acid (OA) and its derivatives—dinophysistoxins—belong to the group of diarrhetic shellfish poison toxins (DSP toxins), which are produced by toxin-forming dinoflagellates. When mollusks ingest these algae, DSP toxins accumulate in their adipose tissue and can cause acute poisoning in humans upon consumption. We developed a simple, a sensitive, and a specific test system for the detection and semiquantitatively determination of okadaic acid and its derivatives in mollusks. This system is based on a direct competitive immunochromatographic analysis. The test strips contain three immunoreagent binding lines: a test line, a control line, and an internal reference line. The color intensity of the internal reference line remains independent of the OA concentration in the solution and is comparable to the color intensity of the test line. If DSP toxins are present in a sample, the color intensity of the test line decreases relative to the reference line, enabling toxin detection. Sample preparation involves the extraction of mollusk tissue homogenates with methanol, followed by the dilution of the extract with an analysis buffer. The test strips are manufactured in a dipstick format, and the analysis procedure consists of immersing part of the strip into a sample for a few seconds, with results processed after 30 min. The total analysis time, including sample preparation, is approximately 1 h. The test system reliably detects and semiquantitatively determines okadaic acid in various mollusks, including oysters, mussels, venus clams, whelks, and sea scallops, at toxin concentrations of up to 40 ng/g using visual result detection and up to 10 ng/g using instrumental registration. These limits of detection are significantly lower than the maximum permissible concentration for okadaic acid set by the World Health Organization and Rospotrebnadzor (160 ng/g). In addition to okadaic acid, the test system can also detect its derivatives, dinophysistoxin-1 and dinophysistoxin-2, in mollusks. The developed immunochromatographic test system can be used for monitoring OA-group toxin levels in mollusks in Russia.