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

Xiong, Hua-Long; Wu, Yang-Tao; Cao, Jia-Li; Yang, Ren; Liu, Ying-Xia; Ma, Jian; Qiao, Xiao-Yang; Yao, Xiang-Yang; Zhang, Bao-Hui; Zhang, Ya-Li; Hou, Wang-Heng; Shi, Yang; Xu, Jing-Jing; Zhang, Liang; 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 neutralization assay is very important for the development of vaccines and novel drugs. In this study, a G protein-deficient vesicular stomatitis virus (VSVdG) bearing a truncated spike protein (S with C-terminal 18 amino acid truncation) was compared to that bearing the full-length spike protein of SARS-CoV-2 and showed much higher efficiency. A neutralization assay was established based on VSV-SARS-CoV-2-Sdel18 pseudovirus and hACE2-overexpressing BHK21 cells (BHK21-hACE2 cells). The experimental results can be obtained by automatically counting the number of EGFP-positive cells at 12 h after infection, making the assay convenient and high-throughput. The serum neutralizing titer measured by the VSV-SARS-CoV-2-Sdel18 pseudovirus assay has a good correlation with that measured by the wild type SARS-CoV-2 assay. Seven neutralizing monoclonal antibodies targeting the receptor binding domain (RBD) of the SARS-CoV-2 S protein were obtained. This efficient and reliable pseudovirus assay model could facilitate the development of new drugs and vaccines.