Liu Jian, Zhu Rui, Ye Xiangzhong, Yang Lianwei, Wang Yongmei, Huang Yanying, Wu Jun, Wang Wei, Ye Jianghui, Li Yimin, Zhao Qinjian, Zhu Hua, Cheng Tong, Xia Ningshao
State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, School of Life Sciences, Xiamen University, Xiamen, 361102, China.
Appl Microbiol Biotechnol. 2015 Jun;99(11):4845-53. doi: 10.1007/s00253-015-6602-5. Epub 2015 May 3.
Varicella-zoster virus (VZV) is a highly infectious agent that causes varicella and herpes zoster (HZ), which may be associated with severe neuralgia. Vaccination is the most effective way to reduce the burden of the diseases. VZV glycoprotein E (gE) is the major and most immunogenic membrane protein that plays important roles in vaccine efficacy. A quantitative assay for gE content is desirable for the VZV vaccine process monitoring and product analysis. In this study, 70 monoclonal antibodies (mAbs) were obtained after immunizing mice with purified recombinant gE (rgE). The collection of mAbs was well-characterized, and a pair of high-affinity neutralization antibodies (capture mAb 4A2 and detection mAb 4H10) was selected to establish a specific and sensitive sandwich enzyme-linked immunosorbent assay (ELISA) to quantify the native and recombinant gE. The detection limit of this assay was found to be 1.95 ng/mL. Furthermore, a reasonably good correlation between the gE content (as measured by the mAb-based quantitative ELISA) and the virus titer (as measured by the "gold standard" plaque assay) was observed when both assays were performed for tracking the kinetics of virus growth during cell culture. A total of 16 batches of lyophilized VZV vaccine were tested using the newly developed quantitative ELISA and classical plaque assay, demonstrating reasonably good correlation between gE content and virus titer. Therefore, this mAb-based gE quantitative assay serves as a rapid, stable, and sensitive method for monitoring viral antigen content, one additional quantitative method for VZV vaccine process and product characterization. This quantitative ELISA may also serve as a complementary method for virus titering.
水痘带状疱疹病毒（VZV）是一种高传染性病原体，可引起水痘和带状疱疹（HZ），后者可能伴有严重神经痛。接种疫苗是减轻这些疾病负担的最有效方法。VZV糖蛋白E（gE）是主要且免疫原性最强的膜蛋白，在疫苗效力中发挥重要作用。对VZV疫苗生产过程监测和产品分析而言，定量检测gE含量很有必要。在本研究中，用纯化的重组gE（rgE）免疫小鼠后获得了70种单克隆抗体（mAb）。对mAb的收集进行了充分表征，并选择了一对高亲和力中和抗体（捕获mAb 4A2和检测mAb 4H10）来建立一种特异且灵敏的夹心酶联免疫吸附测定法（ELISA），以定量天然和重组gE。该测定法的检测限为1.95 ng/mL。此外，在细胞培养过程中进行两种测定以追踪病毒生长动力学时，观察到gE含量（通过基于mAb的定量ELISA测量）与病毒滴度（通过“金标准”蚀斑试验测量）之间存在合理的良好相关性。使用新开发的定量ELISA和经典蚀斑试验对总共16批冻干VZV疫苗进行了检测，结果表明gE含量与病毒滴度之间存在合理的良好相关性。因此，这种基于mAb的gE定量测定法是一种快速、稳定且灵敏的监测病毒抗原含量的方法，是VZV疫苗生产过程和产品特性鉴定的又一种定量方法。这种定量ELISA还可作为病毒滴度测定的补充方法。
