Toward the development of monoclonal antibody-based assays to probe virion-like epitopes in hepatitis B vaccine antigen.
Prophylactic vaccines against hepatitis B Virus (HBV) infection were produced in different expression systems under different processing conditions. Since the recombinant HBV surface antigen (HBsAg) in these vaccines is a cysteine-rich protein with 14 cysteines among a total of 226 amino acids, the epitopes are dependent on the formation of intra- and intermolecular disulfide bonds. A panel of 22 monoclonal antibodies (mAbs) were developed and evaluated with respect to their sensitivity to disulfide reduction treatment of recombinant HBsAg. Not surprisingly, different mAbs showed different degree of sensitivity to controlled HBsAg disulfide reduction. With a view to exploring the functionality of anti-HBsAg mAbs to be used in HBsAg quality analysis, in vitro neutralization activity for the mAbs was assessed. One of the mAbs tested, 5F11, which showed high sensitivity to the disulfide integrity in HBsAg, was shown also to be highly effective in neutralizing HBV in vitro. Conversely, 42B6, while exhibiting similar neutralization activity, showed comparable binding HBsAg with or without reduction treatment. Based on these mAb characteristics, a sandwich ELISA with 42B6 being the capture Ab and detection Ab was developed to quantify HBsAg (like a "mass" assay) during antigen bioprocessing or in vaccine products. In parallel, when 5F11 was used as the detection Ab (with the same capture Ab), the assay can be used to probe disulfide-dependent and virion-like epitopes in intermediates or final products of hepatitis B vaccine, serving as a surrogate marker for vaccine efficacy to elicit neutralizing antibodies. This approach enables the comparative epitope specific antigenicity analysis of HBsAg antigen preparations from different sources.
National Institute of Diagnostics and Vaccine Development in Infectious Diseases; Xiamen University; Xiamen, Fujian, PR China; State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics; Xiamen University; Xiamen, Fujian, PR China; School of Life Sciences; Xiamen University; Xiamen, Fujian, PR China.
Yibin
Y
Zhu
National Institute of Diagnostics and Vaccine Development in Infectious Diseases; Xiamen University; Xiamen, Fujian, PR China; State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics; Xiamen University; Xiamen, Fujian, PR China; School of Life Sciences; Xiamen University; Xiamen, Fujian, PR China.
Tianying
T
Zhang
National Institute of Diagnostics and Vaccine Development in Infectious Diseases; Xiamen University; Xiamen, Fujian, PR China; State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics; Xiamen University; Xiamen, Fujian, PR China; School of Life Sciences; Xiamen University; Xiamen, Fujian, PR China.
Jinghua
J
Zhao
National Institute of Diagnostics and Vaccine Development in Infectious Diseases; Xiamen University; Xiamen, Fujian, PR China; State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics; Xiamen University; Xiamen, Fujian, PR China; School of Life Sciences; Xiamen University; Xiamen, Fujian, PR China.
Zusen
Z
Weng
National Institute of Diagnostics and Vaccine Development in Infectious Diseases; Xiamen University; Xiamen, Fujian, PR China; State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics; Xiamen University; Xiamen, Fujian, PR China; School of Public Health; Xiamen University; Xiamen, Fujian, PR China.
Quan
Q
Yuan
National Institute of Diagnostics and Vaccine Development in Infectious Diseases; Xiamen University; Xiamen, Fujian, PR China; State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics; Xiamen University; Xiamen, Fujian, PR China; School of Life Sciences; Xiamen University; Xiamen, Fujian, PR China; School of Public Health; Xiamen University; Xiamen, Fujian, PR China.
Shaowei
S
Li
National Institute of Diagnostics and Vaccine Development in Infectious Diseases; Xiamen University; Xiamen, Fujian, PR China; State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics; Xiamen University; Xiamen, Fujian, PR China; School of Public Health; Xiamen University; Xiamen, Fujian, PR China.
Jun
J
Zhang
National Institute of Diagnostics and Vaccine Development in Infectious Diseases; Xiamen University; Xiamen, Fujian, PR China; State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics; Xiamen University; Xiamen, Fujian, PR China; School of Life Sciences; Xiamen University; Xiamen, Fujian, PR China; School of Public Health; Xiamen University; Xiamen, Fujian, PR China.
Ningshao
N
Xia
National Institute of Diagnostics and Vaccine Development in Infectious Diseases; Xiamen University; Xiamen, Fujian, PR China; State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics; Xiamen University; Xiamen, Fujian, PR China; School of Public Health; Xiamen University; Xiamen, Fujian, PR China.
Qinjian
Q
Zhao
D000911:Antibodies, Monoclonal; D000956:Antigens, Viral / Q000032:analysis / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D000939:Epitopes / Q000032:analysis / Q000276:immunology*; D006510:Hepatitis B Antibodies; D017325:Hepatitis B Vaccines / Q000276:immunology*; D006801:Humans; D013678:Technology, Pharmaceutical / Q000379:methods*; D064166:Vaccine Potency
D000911:Antibodies, Monoclonal; D000956:Antigens, Viral; D000939:Epitopes; D006510:Hepatitis B Antibodies; D017325:Hepatitis B Vaccines
HBsAg; disulfide; epitope; hepatitis B virus; monoclonal antibody; neutralization activity