Development of cell-based pseudovirus entry assay to identify potential viral entry inhibitors and neutralizing antibodies against SARS-CoV-2

Genes & Diseases
Jie HuKai Wang

Abstract

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 ...Continue Reading

Citations

Feb 2, 2021·Molecular Therapy. Methods & Clinical Development·Xinping FuXiaoliu Zhang
Apr 30, 2021·Journal of Immunological Methods·Aria C Shi, Ping Ren
Apr 27, 2021·Frontiers in Pharmacology·Rui Hong ChenVincent Kam Wai Wong
May 11, 2021·Frontiers in Cellular and Infection Microbiology·Tarun MishraAjit Chande
May 19, 2021·Acta Pharmaceutica Sinica. B·Zinuo ChenQinghua Cui
Jun 29, 2021·Journal of Virological Methods·Peng YangShengqi Wang
Aug 18, 2021·Virologica Sinica·Jiang-Fan LiCheng-Feng Qin

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Datasets Mentioned

BETA
MN908947

Methods Mentioned

BETA
electrophoresis
transfection
Assay
confocal microscopy

Software Mentioned

ImageJ
GraphPad
GraphPad Prism

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