A Novel Blocking ELISA for Detection of Antibodies against Hepatitis E Virus in Domestic Pigs

被引:10
作者
Chen, Yiyang [1 ,2 ]
Zhao, Qin [1 ,2 ]
Liu, Baoyuan [1 ,2 ]
Wang, Lizhen [1 ,2 ]
Sun, Yani [1 ,2 ]
Li, Huixia [1 ,2 ]
Wang, Xinjie [1 ,2 ]
Syed, Shahid Faraz [1 ,2 ]
Zhang, Gaiping [3 ]
Zhou, En-Min [1 ,2 ]
机构
[1] Northwest A&F Univ, Coll Vet Med, Dept Vet Prevent Med, Yangling, Shaanxi, Peoples R China
[2] Minist Agr, Sci Observing & Expt Stn Vet Pharmacol & Vet Biot, Yangling, Shaanxi, Peoples R China
[3] Henan Agr Univ, Coll Anim Sci & Vet Med, Zhengzhou, Henan, Peoples R China
基金
中国国家自然科学基金;
关键词
SEROLOGICAL PREVALENCE; MONOCLONAL-ANTIBODIES; RT-QPCR; CHINA; INFECTION; SWINE; HEV; GENOTYPE; ANIMALS; TRANSMISSION;
D O I
10.1371/journal.pone.0152639
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
Hepatitis E virus (HEV) infects both humans and animals, with an overall human mortality rate generally less than 1%, but as high as 20% among pregnant women. HEV strains fall into 4 major genotypes. Zoonotic genotypes 3 and 4 associate with sporadic human and animal HEV cases in many industrialized countries. To date, collective evidence implicates pigs as the main HEV reservoir, justifying the importance of monitoring HEV infection rates in pig herds to prevent human illness. Due to the lack of a robust in vitro cell culture system for viral propagation, no "gold standard" assay has yet been developed to detect HEV infection in domestic pigs. 1E4, a monoclonal antibody (mAb) specific for the C-terminal 268 amino acids of HEV genotype 4 ORF2 capsid protein (sORF2-C), was generated and conjugated to horseradish peroxidase (HRP) for use in a blocking ELISA (bELISA). Optimal sORF2-C coating antigen concentration (8 mu g/ml), HRP-1E4 dilution (1:1000), and test pig serum dilution (1:20) were determined using a checkerboard titration test. A cut-off value of 16.9% was chosen to differentiate between positive vs. negative sera after mean percent inhibition (PI) testing of 230 negative pig sera. Compared with the indirect ELISA (iELISA), western blot, and a commercial ELISA kit for detecting anti-HEV antibodies in human sera, the bELISA showed no statistical differences and statistically high coincidence of 93.23%, 92%, and 95% with the other tests, respectively. A blocking ELISA (bELISA) for detecting anti-HEV antibodies in pig serum samples was developed with high sensitivity and high specificity comparable to that of the indirect ELISA. The bELISA results exhibited high agreement with iELISA, western blot, and a commercial ELISA kit designed to detect human anti-HEV antibodies. Therefore, bELISA should serve as an ideal method for large-scale serological investigation of anti-HEV antibodies in domestic pigs.
引用
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页数:15
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