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

Shan Zhou; Shengyong Zhang; Mingshu Wang; Anchun Cheng; Dekang Zhu; Shun Chen; Mafeng Liu; Xinxin Zhao; Renyong Jia; Qiao Yang; Ying Wu; Shaqiu Zhang; Yunya Liu; Yanling Yu; Ling Zhang; Xiaoyue Chen

doi:10.1016/j.jviromet.2019.03.012

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

J Virol Methods. 2019 Jun:268:56-61. doi: 10.1016/j.jviromet.2019.03.012. Epub 2019 Mar 21.

Authors

Shan Zhou¹, Shengyong Zhang², Mingshu Wang³, Anchun Cheng⁴, Dekang Zhu⁵, Shun Chen⁶, Mafeng Liu⁷, Xinxin Zhao⁸, Renyong Jia⁹, Qiao Yang¹⁰, Ying Wu¹¹, Shaqiu Zhang¹², Yunya Liu¹³, Yanling Yu¹⁴, Ling Zhang¹⁵, Xiaoyue Chen¹⁶

Affiliations

¹ Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Key Laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Avian Disease Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China. Electronic address: [email protected].
² Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Key Laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Avian Disease Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China. Electronic address: [email protected].
³ Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Key Laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Avian Disease Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China. Electronic address: [email protected].
⁴ Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Key Laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Avian Disease Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China. Electronic address: [email protected].
⁵ Key Laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Avian Disease Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China. Electronic address: [email protected].
⁶ Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Key Laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Avian Disease Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China. Electronic address: [email protected].
⁷ Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Key Laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Avian Disease Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China. Electronic address: [email protected].
⁸ Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Key Laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Avian Disease Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China. Electronic address: [email protected].
⁹ Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Key Laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Avian Disease Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China. Electronic address: [email protected].
¹⁰ Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Key Laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Avian Disease Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China. Electronic address: [email protected].
¹¹ Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Key Laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Avian Disease Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China. Electronic address: [email protected].
¹² Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Key Laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Avian Disease Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China. Electronic address: [email protected].
¹³ Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Key Laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Avian Disease Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China. Electronic address: [email protected].
¹⁴ Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Key Laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Avian Disease Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China. Electronic address: [email protected].
¹⁵ Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Key Laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Avian Disease Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China. Electronic address: [email protected].
¹⁶ Key Laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China; Avian Disease Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu, Sichuan, 611130, PR China. Electronic address: [email protected].

PMID: 30905595
DOI: 10.1016/j.jviromet.2019.03.012

Abstract

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 μg/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.

Keywords: 3A protein; Antibody detection; Duck hepatitis A virus type 1(DHAV-1); Indirect ELISA(I-ELISA).

Publication types

Evaluation Study
Research Support, Non-U.S. Gov't

MeSH terms

Animals
Antibodies, Viral
Cross Reactions
Ducks / immunology*
Enzyme-Linked Immunosorbent Assay*
Escherichia coli / genetics
Hepatitis Antibodies / blood*
Hepatitis Virus, Duck / immunology*
Hepatitis, Viral, Animal / diagnosis*
Hepatitis, Viral, Animal / immunology
Poultry Diseases / diagnosis
Poultry Diseases / virology
Recombinant Proteins / immunology
Sensitivity and Specificity
Viral Proteins / immunology*

Substances

Antibodies, Viral
Hepatitis Antibodies
Recombinant Proteins
Viral Proteins