Establishment, validation and application of fluorescence focus assay for rapid titration of Japanese encephalitis virus
10.3760/cma.j.cn112309-20211109-00374
- VernacularTitle:一种快速检测乙型脑炎病毒滴度的荧光灶试验方法的建立、验证及应用
- Author:
Na LUO
1
;
Jian QIAO
;
Zhijun JIANG
;
Shantang YANG
;
Zhongkai ZHANG
;
Jie WU
;
Gelin XU
Author Information
1. 武汉生物制品研究所有限责任公司轮状病毒疫苗室，武汉 430207
- Keywords:
Live-attenuated Japanese encephalitis vaccine;
Polyclonal antibody;
Fluorescent focus assay;
Plaque assay;
Validation
- From:
Chinese Journal of Microbiology and Immunology
2022;42(3):224-233
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To establish and validate a fluorescence focus assay (FFA) for rapid titration of Japanese encephalitis virus (JEV) and to evaluate its application in the production of Japanese encephalitis vaccine.Methods:Recombinant JEV non-structural protein 1 (NS1) was expressed in a prokaryotic expression system. After purification, JEV-NS1 was used to immunize rabbits to induce polyclonal antibody. FFA was established with the polyclonal antibody to titer JEV. The accuracy of FFA was validated by comparing with plaque assay, and the specificity, precision, linearity, range and robustness of FFA were also validated. Twenty-eight batches of live-attenuated Japanese encephalitis vaccine were titrated with FFA and plaque assay to analyze the relationship between the two assays.Results:FFA established with polyclonal antibody against JEV-NS1 could be used to titrate JEV, and there was no cross reaction with other viruses (tick-borne encephalitis virus, yellow fever virus, coxsackievirus A2, coxsackievirus A4). Results of the validation tests showed that FFA met the requirement of quality control for live-attenuated Japanese encephalitis vaccine. FFA was more consistency than plaque assay.Conclusions:The established FFA could be used for virus titration in the production of live-attenuated Japanese encephalitis vaccine.
