@article{6ad8c15036404ac1b6bd4a2817b19c27,
title = "Protocol to isolate and assess spike protein cleavage in SARS-CoV-2 variants obtained from clinical COVID-19 samples",
abstract = "For efficient cell entry, SARS-CoV-2 spike protein needs to be cleaved by cellular proteases. Here, we present a comprehensive protocol to assess SARS-CoV-2 spike protein cleavage in viral supernatants from SARS-CoV-2-infected cells. We also include a previous step of SARS-CoV-2 isolation from nasopharyngeal swabs of patients with COVID-19. We optimized the procedures to enhance successful viral isolation and specific spike detection. This protocol facilitates the evaluation of the role of spike mutations in spike protein processing. For complete details on the use and execution of this protocol, please refer to Escalera et al. (2022).",
keywords = "Evolutionary biology, Microbiology",
author = "Alba Escalera and Adolfo Garc{\'i}a-Sastre and Teresa Aydillo",
note = "Funding Information: We thank Randy Albrecht for support with the BSL3 facility and procedures at the ISMMS and Richard Cadagan for excellent technical assistance. We also thank Mount Sinai Pathogen Surveillance Program (PSP) for collecting and providing the nasopharyngeal swabs used in this study. This research was partly funded by CRIPT (Center for Research on Influenza Pathogenesis and Transmission), an NIAID funded Center of Excellence for Influenza Research and Response (CEIRR, contract #75N93021C00014) (A.G.-S.), NCI SeroNet grant U54CA260560 (A.G.-S.), NIAID grants U19AI135972 and U19AI142733 (A.G.-S.), DARPA grant HR0011-19-2-0020 (A.G.-S.), DoD grant W81XWH-20-1-0270 (A.G.-S.), JPB Foundation (A.G.-S.), Open Philanthropy Project (research grant 2020-215611 5384) (A.G.-S.), and anonymous donors to A.G.-S. T.A. and A.G.-S. conceived, designed, and supervised research. T.A. provided training to A.E. A.E. performed experiments. A.E. and T.A. analyzed data and wrote the manuscript. A.G.-S. revised the manuscript and acquired funding. The A.G.-S. laboratory has received research support from Pfizer, Senhwa Biosciences, Kenall Manufacturing, Avimex, Johnson & Johnson, Dynavax, 7Hills Pharma, Pharmamar, ImmunityBio, Accurius, Nanocomposix, Hexamer, N-fold LLC, Model Medicines, Atea Pharma, and Merck outside of the reported work. A.G.-S. has consulting agreements for the following companies involving cash and/or stock: Vivaldi Biosciences, Contrafect, 7Hills Pharma, Avimex, Vaxalto, Pagoda, Accurius, Esperovax, Farmak, Applied Biological Laboratories, Pharmamar, Paratus, CureLab Oncology, CureLab Veterinary, Synairgen, and Pfizer outside of the reported work. A.G.-S. has been an invited speaker in meeting events organized by Sequirus, Janssen, and AstraZeneca. A.G.-S. is an inventor on patents and patent applications on the use of antivirals and vaccines for the treatment and prevention of virus infections and cancer, owned by the Icahn School of Medicine at Mount Sinai, New York, outside of the reported work. Funding Information: We thank Randy Albrecht for support with the BSL3 facility and procedures at the ISMMS and Richard Cadagan for excellent technical assistance. We also thank Mount Sinai Pathogen Surveillance Program (PSP) for collecting and providing the nasopharyngeal swabs used in this study. This research was partly funded by CRIPT ( Center for Research on Influenza Pathogenesis and Transmission ), an NIAID funded Center of Excellence for Influenza Research and Response (CEIRR, contract #75N93021C00014 ) (A.G.-S.), NCI SeroNet grant U54CA260560 (A.G.-S.) , NIAID grants U19AI135972 and U19AI142733 (A.G.-S.), DARPA grant HR0011-19-2-0020 (A.G.-S.), DoD grant W81XWH-20-1-0270 (A.G.-S.), JPB Foundation (A.G.-S.), Open Philanthropy Project (research grant 2020-215611 5384 ) (A.G.-S.), and anonymous donors to A.G.-S. Publisher Copyright: {\textcopyright} 2022 The Authors",
year = "2022",
month = sep,
day = "16",
doi = "10.1016/j.xpro.2022.101502",
language = "English",
volume = "3",
journal = "STAR Protocols",
issn = "2666-1667",
publisher = "Cell Press",
number = "3",
}