猪流行性腹泻病毒N蛋白阻断ELISA抗体检测方法的建立及初步应用

doi:10.11843/j.issn.0366-6964.2022.04.017

Abstract

Abstract: The purpose of this study is to establish a blocking ELISA antibodies detection method for porcine epidemic diarrhea virus (PEDV). The purified N protein was used as the coating antigen, and the ELISA reaction conditions were optimized by the chess rboard titration. A blocking ELISA method for detecting PEDV antibodies was established, and its specificity, sensitivity and repeatability tests were carried out. One hundred and forty clinical serum samples were tested, and the results were compared with commercially IDvet PEDV indirect ELISA antibodies detection kit. The results showed that the best antigen coating concentration was 625 ng·mL^-1, and the best dilution ratio of serum was 1:1; The best dilution of the HRP-conjugated antibody working solution was 1:5 000; There was no cross-reaction with healthy pig serum and the positive sera of common pig disease pathogens, such as classical swine fever virus (CSFV), porcine reproductive and respiratory syndrome virus (PRRSV), porcine circovirus type 2 (PCV2), and transmissible gastroenteritis virus (TGEV). The sensitivity of PEDV positive serum was 1:16, which was equivalent to that of IDvet ELISA kit (titer 1∶32). The coefficient of variation of within-run and between-run repeatability test is less than 10%, so it showed that the blocking ELISA established in this study had good repeatability and stability; the kappa value of detected 140 clinical porcine serum using this method was 0.87 when compared with IDvet ELISA. The above results indicated that the established blocking ELISA method for detecting PEDV antibodies in this study could be applied to the prevention and control of PEDV, epidemiological investigation and the monitoring of antibody levels after vaccine immunization.

Key words: porcine epidemic diarrhea variant virus, N protein, antibodies detection, blocking ELISA

CLC Number:

S852.659.6

WAN Ying, ZHOU Gaijing, MA Yuan, SHI Zhengwang, XIAO Shuqi, LUO Juncong, SONG Rui, CAO Liyan, YANG bo, WANG Lijuan, TIAN Hong, ZHENG Haixue. Development and Application of N-protein Blocking ELISA for Detecting Porcine Epidemic Diarrhea Virus Antibodies[J]. Acta Veterinaria et Zootechnica Sinica, 2022, 53(4): 1173-1181.

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Development and Application of N-protein Blocking ELISA for Detecting Porcine Epidemic Diarrhea Virus Antibodies

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