A protease-activatable luminescent biosensor and reporter cell line for authentic SARS-CoV-2 infection.

Autor: Gerber PP; Department of Medicine, University of Cambridge, Cambridge, United Kingdom.; Cambridge Institute for Therapeutic Immunology and Infectious Disease (CITIID), Jeffrey Cheah Biomedical Centre, University of Cambridge, Cambridge, United Kingdom., Duncan LM; Department of Medicine, University of Cambridge, Cambridge, United Kingdom.; Cambridge Institute for Therapeutic Immunology and Infectious Disease (CITIID), Jeffrey Cheah Biomedical Centre, University of Cambridge, Cambridge, United Kingdom., Greenwood EJ; Department of Medicine, University of Cambridge, Cambridge, United Kingdom.; Cambridge Institute for Therapeutic Immunology and Infectious Disease (CITIID), Jeffrey Cheah Biomedical Centre, University of Cambridge, Cambridge, United Kingdom., Marelli S; Department of Medicine, University of Cambridge, Cambridge, United Kingdom.; Cambridge Institute for Therapeutic Immunology and Infectious Disease (CITIID), Jeffrey Cheah Biomedical Centre, University of Cambridge, Cambridge, United Kingdom., Naamati A; Department of Medicine, University of Cambridge, Cambridge, United Kingdom.; Cambridge Institute for Therapeutic Immunology and Infectious Disease (CITIID), Jeffrey Cheah Biomedical Centre, University of Cambridge, Cambridge, United Kingdom., Teixeira-Silva A; Department of Medicine, University of Cambridge, Cambridge, United Kingdom.; Cambridge Institute for Therapeutic Immunology and Infectious Disease (CITIID), Jeffrey Cheah Biomedical Centre, University of Cambridge, Cambridge, United Kingdom., Crozier TW; Department of Medicine, University of Cambridge, Cambridge, United Kingdom.; Cambridge Institute for Therapeutic Immunology and Infectious Disease (CITIID), Jeffrey Cheah Biomedical Centre, University of Cambridge, Cambridge, United Kingdom., Gabaev I; Department of Medicine, University of Cambridge, Cambridge, United Kingdom.; Cambridge Institute for Therapeutic Immunology and Infectious Disease (CITIID), Jeffrey Cheah Biomedical Centre, University of Cambridge, Cambridge, United Kingdom., Zhan JR; Department of Medicine, University of Cambridge, Cambridge, United Kingdom.; Cambridge Institute for Therapeutic Immunology and Infectious Disease (CITIID), Jeffrey Cheah Biomedical Centre, University of Cambridge, Cambridge, United Kingdom., Mulroney TE; MRC Toxicology Unit, University of Cambridge, Cambridge, United Kingdom., Horner EC; MRC Toxicology Unit, University of Cambridge, Cambridge, United Kingdom., Doffinger R; Department of Clinical Biochemistry and Immunology, Cambridge University Hospitals NHS Foundation Trust, Cambridge, United Kingdom., Willis AE; MRC Toxicology Unit, University of Cambridge, Cambridge, United Kingdom., Thaventhiran JE; Department of Medicine, University of Cambridge, Cambridge, United Kingdom.; MRC Toxicology Unit, University of Cambridge, Cambridge, United Kingdom., Protasio AV; Department of Pathology, University of Cambridge, Cambridge, United Kingdom., Matheson NJ; Department of Medicine, University of Cambridge, Cambridge, United Kingdom.; Cambridge Institute for Therapeutic Immunology and Infectious Disease (CITIID), Jeffrey Cheah Biomedical Centre, University of Cambridge, Cambridge, United Kingdom.; NHS Blood and Transplant, Cambridge, United Kingdom.
Jazyk: angličtina
Zdroj: PLoS pathogens [PLoS Pathog] 2022 Feb 10; Vol. 18 (2), pp. e1010265. Date of Electronic Publication: 2022 Feb 10 (Print Publication: 2022).
DOI: 10.1371/journal.ppat.1010265
Abstrakt: Efforts to define serological correlates of protection against COVID-19 have been hampered by the lack of a simple, scalable, standardised assay for SARS-CoV-2 infection and antibody neutralisation. Plaque assays remain the gold standard, but are impractical for high-throughput screening. In this study, we show that expression of viral proteases may be used to quantitate infected cells. Our assays exploit the cleavage of specific oligopeptide linkers, leading to the activation of cell-based optical biosensors. First, we characterise these biosensors using recombinant SARS-CoV-2 proteases. Next, we confirm their ability to detect viral protease expression during replication of authentic virus. Finally, we generate reporter cells stably expressing an optimised luciferase-based biosensor, enabling viral infection to be measured within 24 h in a 96- or 384-well plate format, including variants of concern. We have therefore developed a luminescent SARS-CoV-2 reporter cell line, and demonstrated its utility for the relative quantitation of infectious virus and titration of neutralising antibodies.
Competing Interests: The authors have declared that no competing interests exist.
Databáze: MEDLINE
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