Development of an indirect ELISA method based on the VP3 protein of duck hepatitis A virus type 1 (DHAV-1) for dual detection of DHAV-1 and DHAV-3 antibodies.
An indirect enzyme-linked immunosorbent assay (I-ELISA) based on the recombinant VP3 protein of duck hepatitis A virus type 1 (DHAV-1) was developed and evaluated in this study. The optimal antigen, serum and enzyme-labeled antibody dilutions were 1:160 (0.94μg), 1:160 and 1:2000, respectively. The optimal blocking buffer was 1% gelatin. The cutoff value was determined to be 0.332, and the analytical sensitivity was 1:1280 (OD450-630=0.37). The results of the specificity evaluation showed that no cross-reactivity existed between DHAV-1 antiserum and other common duck-sensitive pathogens, except for duck hepatitis A virus type 3 (DHAV-3), suggesting that this could be a common approach for the simultaneous detection of DHAV-1 and DHAV-3 antibodies. The coefficients of variation (CVs) for all of the tested samples were lower than 10%. The concordance between the I-ELISA based on the VP3 subunit of DHAV-1 and that based on the whole DHAV-1 particle was 96%. These results indicate that the VP3-based I-ELISA method has high sensitivity, specificity, and repeatability and is as effective as the DHAV-1-based I-ELISA method for sero-surveillance. Thus, it may be a convenient and novel method for DHAV antibody detection and epidemiological surveillance of DHAV prevalence.
Avian Diseases Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu City, Sichuan 611130, PR China; Key Laboratory of Animal Diseases and Human Health of Sichuan Province, Wenjiang, Chengdu City, Sichuan 611130, PR China; Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu City, Sichuan 611130, PR China.
Youlin
Y
Shen
Avian Diseases Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu City, Sichuan 611130, PR China; Key Laboratory of Animal Diseases and Human Health of Sichuan Province, Wenjiang, Chengdu City, Sichuan 611130, PR China; Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu City, Sichuan 611130, PR China. Electronic address: chenganchun@vip.163.com.
Anchun
A
Cheng
Avian Diseases Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu City, Sichuan 611130, PR China; Key Laboratory of Animal Diseases and Human Health of Sichuan Province, Wenjiang, Chengdu City, Sichuan 611130, PR China; Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu City, Sichuan 611130, PR China. Electronic address: mshwang@163.com.
Mingshu
M
Wang
Avian Diseases Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu City, Sichuan 611130, PR China; Key Laboratory of Animal Diseases and Human Health of Sichuan Province, Wenjiang, Chengdu City, Sichuan 611130, PR China; Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu City, Sichuan 611130, PR China.
Shun
S
Chen
Avian Diseases Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu City, Sichuan 611130, PR China; Key Laboratory of Animal Diseases and Human Health of Sichuan Province, Wenjiang, Chengdu City, Sichuan 611130, PR China; Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu City, Sichuan 611130, PR China.
Renyong
R
Jia
Key Laboratory of Animal Diseases and Human Health of Sichuan Province, Wenjiang, Chengdu City, Sichuan 611130, PR China; Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu City, Sichuan 611130, PR China.
Dekang
D
Zhu
Avian Diseases Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu City, Sichuan 611130, PR China; Key Laboratory of Animal Diseases and Human Health of Sichuan Province, Wenjiang, Chengdu City, Sichuan 611130, PR China; Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu City, Sichuan 611130, PR China.
Mafeng
M
Liu
Avian Diseases Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu City, Sichuan 611130, PR China; Key Laboratory of Animal Diseases and Human Health of Sichuan Province, Wenjiang, Chengdu City, Sichuan 611130, PR China; Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu City, Sichuan 611130, PR China.
Kunfeng
K
Sun
Avian Diseases Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu City, Sichuan 611130, PR China; Key Laboratory of Animal Diseases and Human Health of Sichuan Province, Wenjiang, Chengdu City, Sichuan 611130, PR China; Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu City, Sichuan 611130, PR China.
Qiao
Q
Yang
Key Laboratory of Animal Diseases and Human Health of Sichuan Province, Wenjiang, Chengdu City, Sichuan 611130, PR China; Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu City, Sichuan 611130, PR China.
Xiaoyue
X
Chen
D000818:Animals; D000956:Antigens, Viral / Q000276:immunology*; D001715:Bird Diseases / Q000175:diagnosis* / Q000821:virology; D003429:Cross Reactions; D004372:Ducks; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D062665:Epidemiological Monitoring; D006508:Hepatitis Antibodies / Q000097:blood*; D004371:Hepatitis Virus, Duck / Q000276:immunology*; D006524:Hepatitis, Viral, Animal / Q000175:diagnosis* / Q000821:virology; D010850:Picornaviridae Infections / Q000175:diagnosis / Q000662:veterinary; D012680:Sensitivity and Specificity; D012698:Serologic Tests / Q000379:methods*; D014764:Viral Proteins / Q000276:immunology*
D000956:Antigens, Viral; D006508:Hepatitis Antibodies; D014764:Viral Proteins
Antibody detection; Duck hepatitis A virus type 1 (DHAV-1); Indirect ELISA (I-ELISA); VP3 protein