Use of the recombinant nonstructural 3A, 3B, and 3AB proteins of foot-and-mouth disease virus in indirect ELISA for differentiation of vaccinated and infected cattle |
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Authors: | A S Yakovleva A V Shcherbakov A V Kan’shina N S Mudrak T A Fomina |
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Institution: | (1) Federal Center for Animal Health, Ministry of Agriculture of the Russian Federation, Vladimir, 600901, Russia |
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Abstract: | Recombinant foot-and-mouth disease virus (FMDV) proteins 3A, 3B, and 3AB were produced by expressing the corresponding genes in Escherichia coli and purified by metal-chelate affinity chromatography. The recombinant proteins were used as antigens in indirect enzyme-linked immunosorbent assay (ELISA) to differentiate between vaccinated and FMD-infected animals. The following parameters were determined: working concentrations of antigens and peroxidase conjugate of cattle anti-IgG, the optimum composition of blocking buffer, and the positive-negative threshold of the reaction. Tests performed with approximately 200 serum samples taken from animals of different immunity states showed that the protocol with protein 3A as the antigen (3A-ELISA) provided the most reliable differentiation. All the newly developed systems proved to outperform the commercial Chekit FMD-3ABC kit in sensitivity, and 3A-ELISA was no less specific. |
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Keywords: | foot-and-mouth disease virus nonstructural proteins expression in Escherichia coli metal-chelate affinity chromatography indirect ELISA diagnostic sensitivity and specificity |
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