非洲猪瘟病毒P30蛋白的表达及抗体液相芯片检测方法的建立

doi:10.11843/j.issn.0366-6964.2023.10.026

Abstract

Abstract: This study was conducted to develop a rapid and accurate method for detection of African swine fever virus (ASFV) antibody based on x-MAP technology. In our study, pET-28a-p30 expression plasmid was constructed, and ASFV recombinant P30 protein was expressed in Escherichia coli. Then the recombinant P30 protein was coupled with carboxylated fluorescent microspheres and an x-MAP multiple bead-based technology was developed for detection of ASFV P30 antibody. The recombinant plasmid was successfully expressed in inclusion body form in Escherichia coli BL21. Western blot analysis showed that recombinant P30 protein could react specifically with ASFV positive serum, indicating that it had good reactivity. The detection threshold of the x-MAP based ASFV antibody detection method is MFI 1 575.7, and the optimal binding protein concentration is 6 μg per 1.25×10⁶ microspheres, the optimal serum dilution is 1:600, and the optimal concentration of secondary antibody is 1 μg·mL^-1, the method has good specificity, sensitivity and repeatability. Ninety-two clinical pig serum samples were tested by x-MAP analysis and compared with the result tested by a commercial ELISA kit. The results showed that the consistency was 92.39%. The recombinant P30 protein was successfully expressed, and the ASFV antibody detection method based on x-MAP technology was established, which provides a new method for the early serological detection of ASFV infection.

Key words: African swine fever virus, P30 protein, prokaryotic expression, liquid chip technology, antibody detection

CLC Number:

S852.659.1

ZHANG Fangyuan, YANG Dawei, QIU Deyang, JIANG Guoqian, LI Guimei, SHAN Hu. Expression of ASFV P30 Protein and Development of ASFV Antibody Detection Method Based on x-MAP Technology[J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(10): 4300-4310.

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Expression of ASFV P30 Protein and Development of ASFV Antibody Detection Method Based on x-MAP Technology

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