Development of cell-based pseudovirus entry assay to identify potential viral entry inhibitors and neutralizing antibodies against SARS-CoV-2
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, ful...
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| Format: | Article |
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KeAi Communications Co., Ltd.
2020-12-01
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| Series: | Genes and Diseases |
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| Online Access: | http://www.sciencedirect.com/science/article/pii/S2352304220300891 |
| _version_ | 1797761250365538304 |
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| author | Jie Hu Qingzhu Gao Changlong He Ailong Huang Ni Tang Kai Wang |
| author_facet | Jie Hu Qingzhu Gao Changlong He Ailong Huang Ni Tang Kai Wang |
| author_sort | Jie Hu |
| collection | DOAJ |
| description | 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. |
| first_indexed | 2024-03-12T19:11:16Z |
| format | Article |
| id | doaj.art-8e5e82bbcdb34cacb2c5302d36f1060e |
| institution | Directory Open Access Journal |
| issn | 2352-3042 |
| language | English |
| last_indexed | 2024-03-12T19:11:16Z |
| publishDate | 2020-12-01 |
| publisher | KeAi Communications Co., Ltd. |
| record_format | Article |
| series | Genes and Diseases |
| spelling | doaj.art-8e5e82bbcdb34cacb2c5302d36f1060e2023-08-02T05:55:27ZengKeAi Communications Co., Ltd.Genes and Diseases2352-30422020-12-0174551557Development of cell-based pseudovirus entry assay to identify potential viral entry inhibitors and neutralizing antibodies against SARS-CoV-2Jie Hu0Qingzhu Gao1Changlong He2Ailong Huang3Ni Tang4Kai Wang5Key Laboratory of Molecular Biology for Infectious Diseases (Ministry of Education), Institute for Viral Hepatitis, Department of Infectious Diseases, The Second Affiliated Hospital, Chongqing Medical University, Chongqing, 400010, ChinaKey Laboratory of Molecular Biology for Infectious Diseases (Ministry of Education), Institute for Viral Hepatitis, Department of Infectious Diseases, The Second Affiliated Hospital, Chongqing Medical University, Chongqing, 400010, ChinaKey Laboratory of Molecular Biology for Infectious Diseases (Ministry of Education), Institute for Viral Hepatitis, Department of Infectious Diseases, The Second Affiliated Hospital, Chongqing Medical University, Chongqing, 400010, ChinaKey Laboratory of Molecular Biology for Infectious Diseases (Ministry of Education), Institute for Viral Hepatitis, Department of Infectious Diseases, The Second Affiliated Hospital, Chongqing Medical University, Chongqing, 400010, ChinaCorresponding author.; Key Laboratory of Molecular Biology for Infectious Diseases (Ministry of Education), Institute for Viral Hepatitis, Department of Infectious Diseases, The Second Affiliated Hospital, Chongqing Medical University, Chongqing, 400010, ChinaCorresponding author.; Key Laboratory of Molecular Biology for Infectious Diseases (Ministry of Education), Institute for Viral Hepatitis, Department of Infectious Diseases, The Second Affiliated Hospital, Chongqing Medical University, Chongqing, 400010, ChinaSevere 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.http://www.sciencedirect.com/science/article/pii/S2352304220300891Antiviral therapeuticsCoronavirusNeutralizing antibodiesPseudovirusSARS-CoV-2Spike protein |
| spellingShingle | Jie Hu Qingzhu Gao Changlong He Ailong Huang Ni Tang Kai Wang Development of cell-based pseudovirus entry assay to identify potential viral entry inhibitors and neutralizing antibodies against SARS-CoV-2 Genes and Diseases Antiviral therapeutics Coronavirus Neutralizing antibodies Pseudovirus SARS-CoV-2 Spike protein |
| title | Development of cell-based pseudovirus entry assay to identify potential viral entry inhibitors and neutralizing antibodies against SARS-CoV-2 |
| title_full | Development of cell-based pseudovirus entry assay to identify potential viral entry inhibitors and neutralizing antibodies against SARS-CoV-2 |
| title_fullStr | Development of cell-based pseudovirus entry assay to identify potential viral entry inhibitors and neutralizing antibodies against SARS-CoV-2 |
| title_full_unstemmed | Development of cell-based pseudovirus entry assay to identify potential viral entry inhibitors and neutralizing antibodies against SARS-CoV-2 |
| title_short | Development of cell-based pseudovirus entry assay to identify potential viral entry inhibitors and neutralizing antibodies against SARS-CoV-2 |
| title_sort | development of cell based pseudovirus entry assay to identify potential viral entry inhibitors and neutralizing antibodies against sars cov 2 |
| topic | Antiviral therapeutics Coronavirus Neutralizing antibodies Pseudovirus SARS-CoV-2 Spike protein |
| url | http://www.sciencedirect.com/science/article/pii/S2352304220300891 |
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