title: Development of a blocking ELISA for detection of antibodies against avian hepatitis E virus.
abstract: A blocking enzyme-linked immunosorbent assay (bELISA) was developed for the detection of immunoglobulin G antibodies against avian hepatitis E virus (HEV). In the bELISA, the coating antigen was a truncated protein containing C-terminal 268-amino acid region of ORF2 from an avian HEV strain isolated in China (CaHEV) and blocking antibody was a monoclonal antibody (mAb) 1H5 recognizing the epitope within amino acids 384-414 in the C-terminal 268-amino acid region. The concentration of blocking mAb 1H5 was determined as that yielded an OD450nm value of 1.0 for binding to the coating antigen and the antigen concentration and serum dilution were optimized using a checkerboard titration. A cut-off value of 20.7% at the mean percent inhibition plus 3 standard deviations was determined by testing 265 negative sera. The bELISA had a sensitivity of 98.3% by testing 116 positive sera from chickens infected experimentally with CaHEV and had no cross-reaction with other anti-avian virus antibodies. The compliance rates of the bELISA with indirect ELISA and Western blot were 83.7% and 93.3%, respectively, by testing 300 field chicken sera. These results suggested that the bELISA developed in this study can be used for detection of antibodies against avian HEV and showed high reproducibility compared with indirect ELISA and Western blot methods.
authors.affiliation: Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi 712100, China; Experimental Station of Veterinary Pharmacology and Veterinary Biotechnology, China Ministry of Agriculture, Yangling, Shaanxi 712100, China.
authors.forename: Baoyuan
authors.initials: B
authors.lastname: Liu
authors.affiliation: Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi 712100, China; Experimental Station of Veterinary Pharmacology and Veterinary Biotechnology, China Ministry of Agriculture, Yangling, Shaanxi 712100, China.
authors.forename: Qin
authors.initials: Q
authors.lastname: Zhao
authors.affiliation: Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi 712100, China; Experimental Station of Veterinary Pharmacology and Veterinary Biotechnology, China Ministry of Agriculture, Yangling, Shaanxi 712100, China.
authors.forename: Yani
authors.initials: Y
authors.lastname: Sun
authors.affiliation: Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi 712100, China; Experimental Station of Veterinary Pharmacology and Veterinary Biotechnology, China Ministry of Agriculture, Yangling, Shaanxi 712100, China.
authors.forename: Xinjie
authors.initials: X
authors.lastname: Wang
authors.affiliation: Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi 712100, China; Experimental Station of Veterinary Pharmacology and Veterinary Biotechnology, China Ministry of Agriculture, Yangling, Shaanxi 712100, China.
authors.forename: Jinan
authors.initials: J
authors.lastname: Zhao
authors.affiliation: Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi 712100, China; Experimental Station of Veterinary Pharmacology and Veterinary Biotechnology, China Ministry of Agriculture, Yangling, Shaanxi 712100, China.
authors.forename: Taofeng
authors.initials: T
authors.lastname: Du
authors.affiliation: Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi 712100, China; Experimental Station of Veterinary Pharmacology and Veterinary Biotechnology, China Ministry of Agriculture, Yangling, Shaanxi 712100, China.
authors.forename: Chengbao
authors.initials: C
authors.lastname: Wang
authors.affiliation: Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi 712100, China; Experimental Station of Veterinary Pharmacology and Veterinary Biotechnology, China Ministry of Agriculture, Yangling, Shaanxi 712100, China.
authors.forename: Shuqi
authors.initials: S
authors.lastname: Xiao
authors.affiliation: Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi 712100, China; Experimental Station of Veterinary Pharmacology and Veterinary Biotechnology, China Ministry of Agriculture, Yangling, Shaanxi 712100, China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Mu
authors.affiliation: Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi 712100, China; Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Henan Agricultural University, Zhengzhou, Henan 450002, China.
authors.forename: Gaiping
authors.initials: G
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Veterinary Etiological Biology, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, Gansu 730046, China.
authors.forename: Jianxun
authors.initials: J
authors.lastname: Luo
authors.affiliation: Department of Biomedical Sciences, College of Veterinary Medicine, Iowa State University, Ames, IA 50011, USA.
authors.forename: Walter H
authors.initials: WH
authors.lastname: Hsu
authors.affiliation: Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi 712100, China; Experimental Station of Veterinary Pharmacology and Veterinary Biotechnology, China Ministry of Agriculture, Yangling, Shaanxi 712100, China. Electronic address: zhouem@nwsuaf.edu.cn.
authors.forename: En-Min
authors.initials: EM
authors.lastname: Zhou
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000097:blood*; D002645:Chickens; D002681:China; D003429:Cross Reactions; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D006524:Hepatitis, Viral, Animal / Q000175:diagnosis* / Q000821:virology; D052678:Hepevirus / Q000276:immunology*; D011201:Poultry Diseases / Q000175:diagnosis* / Q000821:virology; D012327:RNA Virus Infections / Q000175:diagnosis / Q000662:veterinary* / Q000821:virology; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
chemical_list: D000914:Antibodies, Viral
keywords: Avian hepatitis E virus; Blocking ELISA; Monoclonal antibody; ORF2 protein