Development and evaluation of an indirect ELISA based on recombinant nonstructural protein 3A to detect antibodies to duck hepatitis A virus type 1

被引:12
作者
Zhou, Shan [1 ,2 ,3 ]
Zhang, Shengyong [1 ,2 ,3 ]
Wang, Mingshu [1 ,2 ,3 ]
Cheng, Anchun [1 ,2 ,3 ]
Zhu, Dekang [2 ,3 ]
Chen, Shun [1 ,2 ,3 ]
Liu, Mafeng [1 ,2 ,3 ]
Zhao, Xinxin [1 ,2 ,3 ]
Jia, Renyong [1 ,2 ,3 ]
Yang, Qiao [1 ,2 ,3 ]
Wu, Ying [1 ,2 ,3 ]
Zhang, Shaqiu [1 ,2 ,3 ]
Liu, Yunya [1 ,2 ,3 ]
Yu, Yanling [1 ,2 ,3 ]
Zhang, Ling [1 ,2 ,3 ]
Chen, Xiaoyue [2 ,3 ]
机构
[1] Sichuan Agr Univ, Inst Prevent Vet Med, Chengdu 611130, Sichuan, Peoples R China
[2] Sichuan Agr Univ, Key Lab Anim Dis & Human Hlth Sichuan Prov, Chengdu 611130, Sichuan, Peoples R China
[3] Sichuan Agr Univ, Coll Vet Med, Avian Dis Res Ctr, Chengdu 611130, Sichuan, Peoples R China
关键词
Duck hepatitis A virus type 1(DHAV-1); 3A protein; Indirect ELISA(I-ELISA); Antibody detection; IMMUNOSORBENT-ASSAY ELISA; DIFFERENTIATE ANTIBODIES; RESPIRATORY SYNDROME; SEROLOGIC DETECTION; ENTERITIS VIRUS; VP1; EXPRESSION; INFECTION;
D O I
10.1016/j.jviromet.2019.03.012
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
To develop an indirect enzyme-linked immunosorbent assay(I-ELISA) method based on 3A protein of duck hepatitis A virus type 1(DHAV-1) for detection of DHAV-1 antibody, the recombinant protein 3A of DHAV-1 was expressed in E.coli and detected by Western blotting with DHAV-1 infected duck serum. A 3A-ELISA method using the expressed 3A protein as coating antigen for the detection of antibodies to DHAV-1 was developed. The optimal antigen, serum and enzyme-labeled antibody dilutions were 1:200(6.185 mu g/ml), 1:20 and 1:2000, respectively. The optimal blocking buffer was 5% BSA. The cutoff value was determined to be 0.274, and the analytical sensitivity was 1:1280. There was no cross reaction between DHAV-1 infected duck serum and other common pathogenic duck serum, indicating that I-ELISA could be used to detect DHAV-1 infected duck serum. The coefficients of variation(CVs) were lower than 10%. The concordance between the I-ELISA based on the 3A subunit of DHAV-1 and that based on the whole DHAV-1 particle was 92.7%. Taken together, the 3A-ELISA method is a highly sensitive and specific test that could be used for screening for DHAV-1 infection and monitoring DHAV-1 antibody.
引用
收藏
页码:56 / 61
页数:6
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