Development and validation of an immunochromatographic test system for the determination of okadaic acid in seafood

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Okadaic acid (OA) and its derivatives, dinophysistoxins, belong to the group of diarrhoeal toxins (DSP-toxins) produced by toxin-producing dinoflagellates. When algae are ingested by molluscs, DSP-toxins concentrate in the fatty tissue of the animals and can lead to acute poisoning in humans if consumed. We have developed a simple, sensitive and specific test system for the detection and semi-quantitative rapid determination of OA and its derivatives in shellfish, based on a direct competitive immunochromatographic assay. The test strips have three immunoreagent binding lines – test line, control line and internal comparison line, the staining intensity of which does not depend on the concentration of OA in the solution and is comparable to the staining intensity of the test line. When DSP toxins are present in the sample, the staining intensity of the test line decreases relative to the reference line, which allows detection of the presence of toxins in the samples. Sample preparation includes extraction of mollusk tissue homogenates with methanol followed by dilution of the extract with the solution for analysis. The test strips are manufactured in a “dip-stick” format and the analysis procedure involves dipping part of the test strip into the sample for a few seconds with results processed after 30 min. The total duration of the analysis including sample preparation is about 1 h. The test system allows reliable detection and semi-quantitative determination of OA in various shellfish (oysters, mussels, vongole, Buccinidae, scallops) with the toxin content not more than 40 ng/g (visual method of results registration) and not more than 10 ng/g (instrumental method of results registration), which is significantly lower than the maximum permissible concentration for OA established by the World Health Organisation and Rospotrebnadzor (160 ng/g). In addition to OA, the test system is able to detect its derivatives, dinophysistoxin-1 and dinophysistoxin-2, in molluscs. The developed immunochromatographic test system can be used for monitoring the content of OA-group toxins in molluscs inhabiting Russia.

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作者简介

E. Singov

Federal State Unitary Enterprise “Scientific Center “Signal”

Email: v_vrublevskaya@mail.ru
俄罗斯联邦, Moscow

O. Morenkov

Institute of Cell Biophysics, Federal Research Center “Pushchino Scientific Center for Biological Research of the Russian Academy of Sciences”

Email: v_vrublevskaya@mail.ru
俄罗斯联邦, Pushchino

S. Sipin

Federal State Unitary Enterprise “Scientific Center “Signal”

Email: v_vrublevskaya@mail.ru
俄罗斯联邦, Moscow

V. Vrublevskaya

Institute of Cell Biophysics, Federal Research Center “Pushchino Scientific Center for Biological Research of the Russian Academy of Sciences”

编辑信件的主要联系方式.
Email: v_vrublevskaya@mail.ru
俄罗斯联邦, Pushchino

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2. Fig. 1. Chemical structure of okadaic acid and dinophysistoxins.

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3. Fig. 2. Design of the immunochromatographic test strip and the analysis procedure.

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4. Fig. 3. Detection of okadaic acid in mollusc tissues using a test system with visual detection.

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5. Fig. 4. Dependence of the inhibition value of the test line staining intensity on the okadaic acid content (a) in the physiological solution and the mussel tissue homogenate extract, (b) in the mussel tissue homogenate. Each point represents the arithmetic mean ± standard deviation (n = 5).

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6. Fig. 5. Repeatability of results in the analysis of mussel tissue homogenate. (a) – okadaic acid is absent, (b) – okadaic acid content in mussel tissue homogenate is 40 ng/g, (c) – okadaic acid content in mussel tissue homogenate is 160 ng/g (established MAC level).

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