Establishment and evaluation of a triple-color human papillomavirus pseudovirion neutralization assay
10.3760/cma.j.issn.0253-9624.2018.10.014
- VernacularTitle: 三色混合荧光HPV假病毒中和检测系统的建立及评价
- Author:
Shuangping WEI
1
;
Fei FAN
;
Jie CHEN
;
Xinlin LIU
;
Yurou YANG
;
Zhiping WANG
;
Shuo SONG
;
Zhihai LI
;
Minxi WEI
;
Daning WANG
;
Shaowei LI
;
Ningshao XIA
Author Information
1. National Institute of Diagnostics and Vaccine Development of Infectious Diseases, State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, School of Life Sciences, Xiamen University, Xiamen 361102, China
- Publication Type:Journal Article
- Keywords:
Vaccine;
Neutralization tests;
Immune sera;
Human papillomavirus;
HPV triple-color pseudovirus
- From:
Chinese Journal of Preventive Medicine
2018;52(10):1039-1044
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To establish a triple-color pseudovirion-based neutralization assay (PBNA) and evaluate its capability of detecting immunogenicity of the sera generated by the immunization of HPV 9-valent vaccine.
Methods:HPV pseudovirus (PsVs) 6/11/16/18/31/33/45/52/58 with the encapsidated fluorescence expressing red fluorescent plasmid N31-MCHREEY, green fluorescent N31-EGFP or blue fluorescent N31-mTagBFP were generated. The concentration of HPV PsVs and the infection titers of HPV PsVs were detected by double-antibody sandwich ELISA and TCID50, respectively. The single- and triple color HPV 16/33/45 PsVs were used to detect the neutralization titers of mice sera immunized with HPV 9-valent vaccine and confirmed the accuracy and specificity of the triple-color PBNAs. Then, the single- and triple color HPV 6/11/18/31/33/45/52/58 PsVs were employed to detect the neutralization titers of cynomolgus macaques sera immunized with HPV 9-valent vaccine and determined whether the triple-color PBNAs could be applied to evaluate the immunogenicity of the sera generated by the immunization of HPV9-valent vaccine.
Results:The concentration of HPV16 PsVs encapsulating green, red or blue fluorescent plasmid was 5.0 to 6.0 μg/ml and HPV6/11/18/31/33/45/52/59 triple-color HPV PsVs was about 1.0 to 3.0 μg/ml. 9 types HPV PsVs containing EGFP, Mcherry or mTagBFP reporter plasmid were obtained and the concentration can meet the need of neutralization detection. 9 types single-color fluorescent HPV PsVs had similar infectivity against 293FT cells with the infection titer values between 1×104 and 1×105. The results of PBNAs showed that there was no significant difference in the anti-HPV neutralization titers of mice sera induced by HPV 9-valent vaccine between single-color and triple-color HPV16/33/45 PsVs (P>0.05). Similarly, there was also no significant difference in the anti-HPV neutralization titers of cynomolgus macaques sera induced by HPV 9-valent vaccine between single-color and triple-color HPV6/11/18/31/33/45/52/58 PsVs (P>0.05).
Conclusion:We successfully established the triple-color PBNAs and verified the accuracy and specificity of triple-color PBNAs consistent with single-color PBNAs. The triple-color PBNAs can be applied to evaluate the immunogenicity of HPV 9-valent vaccine's immune serum.
