A high-throughput neutralizing assay for antibodies and sera against hepatitis E virus.
Hepatitis E virus (HEV) is the aetiological agent of enterically transmitted hepatitis. The traditional methods for evaluating neutralizing antibody titres against HEV are real-time PCR and the immunofluorescence foci assay (IFA), which are poorly repeatable and operationally complicated, factors that limit their applicability to high-throughput assays. In this study, we developed a novel high-throughput neutralizing assay based on biotin-conjugated p239 (HEV recombinant capsid proteins, a.a. 368-606) and staining with allophycocyanin-conjugated streptavidin (streptavidin APC) to amplify the fluorescence signal. A linear regression analysis indicated that there was a high degree of correlation between IFA and the novel assay. Using this method, we quantitatively evaluated the neutralization of sera from HEV-infected and vaccinated macaques. The anti-HEV IgG level had good concordance with the neutralizing titres of macaque sera. However, the neutralization titres of the sera were also influenced by anti-HEV IgM responses. Further analysis also indicated that, although vaccination with HEV vaccine stimulated higher anti-HEV IgG and neutralization titres than infection with HEV in macaques, the proportions of neutralizing antibodies in the infected macaques' sera were higher than in the vaccinated macaques with the same anti-HEV IgG levels. Thus, the infection more efficiently stimulated neutralizing antibody responses.
State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, National Institute of Diagnostics and Vaccine Development in Infectious Diseases, School of Life Sciences, Xiamen University, Xiamen, Fujian 361005, PR China.
Wei
W
Cai
State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, National Institute of Diagnostics and Vaccine Development in Infectious Diseases, School of Public Health, Xiamen University, Xiamen, Fujian 361005, PR China.
Zi-Min
ZM
Tang
State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, National Institute of Diagnostics and Vaccine Development in Infectious Diseases, School of Public Health, Xiamen University, Xiamen, Fujian 361005, PR China.
Gui-Ping
GP
Wen
State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, National Institute of Diagnostics and Vaccine Development in Infectious Diseases, School of Public Health, Xiamen University, Xiamen, Fujian 361005, PR China.
Si-Ling
SL
Wang
State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, National Institute of Diagnostics and Vaccine Development in Infectious Diseases, School of Life Sciences, Xiamen University, Xiamen, Fujian 361005, PR China.
Wen-Fang
WF
Ji
State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, National Institute of Diagnostics and Vaccine Development in Infectious Diseases, School of Life Sciences, Xiamen University, Xiamen, Fujian 361005, PR China.
Min
M
Yang
State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, National Institute of Diagnostics and Vaccine Development in Infectious Diseases, School of Life Sciences, Xiamen University, Xiamen, Fujian 361005, PR China.
Dong
D
Ying
State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, National Institute of Diagnostics and Vaccine Development in Infectious Diseases, School of Public Health, Xiamen University, Xiamen, Fujian 361005, PR China.
Zi-Zheng
ZZ
Zheng
State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, National Institute of Diagnostics and Vaccine Development in Infectious Diseases, School of Life Sciences, Xiamen University, Xiamen, Fujian 361005, PR China.|State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, National Institute of Diagnostics and Vaccine Development in Infectious Diseases, School of Public Health, Xiamen University, Xiamen, Fujian 361005, PR China.
Ning-Shao
NS
Xia
D000818:Animals; D057134:Antibodies, Neutralizing / Q000032:analysis*; D000914:Antibodies, Viral / Q000032:analysis*; D036022:Capsid Proteins / Q000276:immunology*; D056945:Hep G2 Cells; D016751:Hepatitis E / Q000517:prevention & control / Q000821:virology; D016752:Hepatitis E virus / Q000276:immunology / Q000378:metabolism*; D057166:High-Throughput Screening Assays / Q000379:methods; D006801:Humans; D008251:Macaca / Q000276:immunology / Q000821:virology; D009500:Neutralization Tests / Q000379:methods*; D014611:Vaccination
D057134:Antibodies, Neutralizing; D000914:Antibodies, Viral; D036022:Capsid Proteins