Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative virus of the coronavirus disease 2019 (COVID-19) pandemic. To establish a safe and convenient assay system for studying entry inhibitors and neutralizing antibodies against SARS-CoV-2, we constructed a codon-optimized, full-length C-terminal mutant spike (S) gene of SARS-CoV-2. We generated a luciferase (Luc)-expressing pseudovirus containing the wild-type or mutant S protein of SARS-CoV-2 in the envelope-defective HIV-1 backbone. The key parameters for this pseudovirus-based assay, including the S mutants and virus incubation time, were optimized. This pseudovirus contains a Luc reporter gene that enabled us to easily quantify virus entry into angiotensin-converting enzyme 2 (ACE2)-expressing 293T cells. Cathepsin (Cat)B/L inhibitor E−64d could significantly block SARS-CoV-2 pseudovirus infection in 293T-ACE2 cells. Furthermore, the SARS-CoV-2 spike pseudotyped virus could be neutralized by sera from convalescent COVID-19 patients or recombinant ACE2 with the fused Fc region of human IgG1. Thus, we developed a pseudovirus-based assay for SARS-CoV-2, which will be valuable for evaluating viral entry inhibitors and neutralizing antibodies against this highly pathogenic virus.

严重急性呼吸综合症冠状病毒2（SARS-CoV-2）是2019年冠状病毒病（COVID-19）大流行的致病病毒。为了建立一个安全方便的测定系统来研究进入抑制剂和中和针对SARS-CoV-2的抗体，我们构建了一个经过密码子优化的SARS-CoV-2全长C末端突变体加标（S）基因。我们生成了一种表达荧光素酶（Luc）的假病毒，在包膜缺陷性HIV-1主链中包含SARS-CoV-2的野生型或突变S蛋白。优化了基于伪病毒的分析的关键参数，包括S突变体和病毒孵育时间。该伪病毒包含一个Luc报告基因使我们能够轻松量化病毒进入表达血管紧张素转化酶2（ACE2）的293T细胞的数量。组织蛋白酶（Cat）B / L抑制剂E-64d可以显着阻断293T-ACE2细胞中的SARS-CoV-2假病毒感染。此外，SARS-CoV-2穗型假型病毒可以被恢复期COVID-19患者的血清或具有人IgG1融合Fc区的重组ACE2的血清中和。因此，我们开发了一种基于伪病毒的SARS-CoV-2检测方法，这对于评估病毒进入抑制剂和中和针对这种高致病性病毒的抗体非常有价值。