Display options
Cite
Barrow KA, Rich LM, Vanderwall ER, et al. Inactivation of Material from SARS-CoV-2-Infected Primary Airway Epithelial Cell Cultures. Methods Protoc. 2021;4(1)doi: 10.3390/mps4010007.
Barrow, K. A., Rich, L. M., Vanderwall, E. R., Reeves, S. R., Rathe, J. A., White, M. P., & Debley, J. S. (2021). Inactivation of Material from SARS-CoV-2-Infected Primary Airway Epithelial Cell Cultures. Methods and protocols, 4(1), . https://doi.org/10.3390/mps4010007
Barrow, Kaitlyn A, et al. "Inactivation of Material from SARS-CoV-2-Infected Primary Airway Epithelial Cell Cultures." Methods and protocols vol. 4,1 (2021). doi: https://doi.org/10.3390/mps4010007
Barrow KA, Rich LM, Vanderwall ER, Reeves SR, Rathe JA, White MP, Debley JS. Inactivation of Material from SARS-CoV-2-Infected Primary Airway Epithelial Cell Cultures. Methods Protoc. 2021 Jan 07;4(1). doi: 10.3390/mps4010007. PMID: 33430421; PMCID: PMC7839057.
Copy Download .nbib Format: NLM
Share it on

Methods Protoc. 2021 Jan 07;4(1). doi: 10.3390/mps4010007.

Inactivation of Material from SARS-CoV-2-Infected Primary Airway Epithelial Cell Cultures.

Methods and protocols

Kaitlyn A Barrow, Lucille M Rich, Elizabeth R Vanderwall, Stephen R Reeves, Jennifer A Rathe, Maria P White, Jason S Debley

Affiliations

  1. Center for Immunity and Immunotherapies, Seattle Children's Research Institute, Seattle, WA 98101, USA.
  2. Department of Pediatrics, Division of Pulmonary and Sleep Medicine, Seattle Children's Hospital, University of Washington, Seattle, WA 98101, USA.
  3. Department of Pediatrics, Division of Infectious Disease, Seattle Children's Hospital, University of Washington, Seattle, WA 98101, USA.

PMID: 33430421 PMCID: PMC7839057 DOI: 10.3390/mps4010007

Abstract

Given that the airway epithelium is the initial site of infection, study of primary human airway epithelial cells (AEC) infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) will be crucial to improved understanding of viral entry factors and innate immune responses to the virus. Centers for Disease Control and Prevention (CDC) guidance recommends work with live SARS-CoV-2 in cell culture be conducted in a Biosafety Level 3 (BSL-3) laboratory. To facilitate downstream assays of materials from experiments there is a need for validated protocols for SARS-CoV-2 inactivation to facilitate safe transfer of material out of a BSL-3 laboratory. We propagated stocks of SARS-CoV-2, then evaluated the effectiveness of heat (65 °C) or ultraviolet (UV) light inactivation. We infected differentiated human primary AECs with SARS-CoV-2, then tested protocols designed to inactivate SARS-CoV-2 in supernatant, protein isolate, RNA, and cells fixed for immunohistochemistry by exposing Vero E6 cells to materials isolated/treated using these protocols. Heating to 65 °C for 10 min or exposing to UV light fully inactivated SARS-CoV-2. Furthermore, we found in SARS-CoV-2-infected primary AEC cultures that treatment of supernatant with UV light, isolation of RNA with Trizol

Keywords: COVID-19; RNA; SARS-CoV-2; Vero E6; airway; epithelial; heat; inactivation; protein

References

  1. Viruses. 2020 Jun 08;12(6): - PubMed
  2. J Allergy Clin Immunol. 2012 Apr;129(4):990-7.e6 - PubMed
  3. Virology. 2020 Sep;548:39-48 - PubMed
  4. J Infect Dis. 2020 Oct 1;222(9):1462-1467 - PubMed
  5. Inhal Toxicol. 2013 Oct;25(12):653-60 - PubMed
  6. Viruses. 2020 Jul 07;12(7): - PubMed
  7. J Med Virol. 2020 Jun;92(6):577-583 - PubMed
  8. Proc Natl Acad Sci U S A. 2020 Sep 29;117(39):24450-24458 - PubMed
  9. J Clin Virol. 2020 Sep;130:104579 - PubMed
  10. J Microbiol Immunol Infect. 2020 Jun;53(3):404-412 - PubMed
  11. J Clin Microbiol. 2020 Oct 21;58(11): - PubMed
  12. MMWR Morb Mortal Wkly Rep. 2020 Mar 27;69(12):343-346 - PubMed
  13. Respir Res. 2015 Feb 13;16:21 - PubMed
  14. J Allergy Clin Immunol. 2014 Sep;134(3):663-670.e1 - PubMed
  15. J Allergy Clin Immunol. 2012 Nov;130(5):1187-1196.e5 - PubMed
  16. J Med Virol. 2020 Jun;92(6):568-576 - PubMed
  17. N Engl J Med. 2003 May 15;348(20):1953-66 - PubMed
  18. Infect Dis Health. 2021 Feb;26(1):63-66 - PubMed
  19. Nat Commun. 2020 Sep 23;11(1):4812 - PubMed
  20. J Allergy Clin Immunol. 2018 Aug;142(2):451-459 - PubMed
  21. Am J Infect Control. 2020 Sep 4;: - PubMed

Publication Types

Grant support