Robust neutralization assay based on SARS-CoV-2 S-bearing vesicular stomatitis virus (VSV) pseudovirus and ACE2-overexpressed BHK21 cells

Xiong, Hua-Long; Wu, Yang-Tao; Cao, Jia-Li; Yang, Ren; Ma, Jian; Qiao, Xiao-Yang; Yao, Xiang-Yang; Zhang, Bao-Hui; Zhang, Ya-Li; Hou, Wang-Heng; Yang-Shi,; Xu, Jing-Jing; Liang-Zhang,; Wang, Shao-Juan; Fu, Bao-Rong; Yang, Ting; Ge, Sheng-Xiang; Zhang, Jun; Yuan, Quan; Huang, Bao-Ying; Li, Zhi-Yong; Zhang, Tian-Ying; Xia, Ning-Shao

The global pandemic of Coronavirus disease 2019 (COVID-19) is a disaster for human society. A convenient and reliable in vitro neutralization assay is very important for the development of neutralizing antibodies, vaccines and other inhibitors. In this study, G protein-deficient vesicular stomatitis virus (VSVdG) bearing full-length and truncated spike (S) protein of SARS-CoV-2 were evaluated. The virus packaging efficiency of VSV-SARS-CoV-2-Sdel18 (S with C-terminal 18 amino acid truncation) is much higher than VSV-SARS-CoV-2-S. A neutralization assay for antibody screening and serum neutralizing titer quantification was established based on VSV-SARS-CoV-2-Sdel18 pseudovirus and human angiotensin-converting enzyme 2 (ACE2) overexpressed BHK21 cell (BHK21-hACE2). The experimental results can be obtained by automatically counting EGFP positive cell number at 12 hours after infection, making the assay convenient and high-throughput. The serum neutralizing titer of COVID-19 convalescent patients measured by VSV-SARS-CoV-2-Sdel18 pseudovirus assay has a good correlation with live SARS-CoV-2 assay. Seven neutralizing monoclonal antibodies targeting receptor binding domain (RBD) of SARS-CoV-2-S were obtained. This efficient and reliable pseudovirus assay model could facilitate the development of new drugs and vaccines.