Essential Site Maintenance: Authorea-powered sites will be updated circa 15:00-17:00 Eastern on Tuesday 5 November.
There should be no interruption to normal services, but please contact us at [email protected] in case you face any issues.

loading page

Production of Trimeric SARS-CoV-2 Spike Protein by CHO Cells for Serological COVID-19 Testing
  • +12
  • Yusuf Johari,
  • Stephen Jaffe,
  • Joseph Scarrott,
  • Abayomi Johnson,
  • ThĂ©o Mozzanino,
  • Thilo Pohle,
  • Sheetal Maisuria,
  • Amina Bhayat-Cammack,
  • Adam Brown,
  • Kang Lan Tee,
  • Philip Jackson,
  • Tuck Seng Wong,
  • Mark Dickman,
  • Ravishankar Sargur,
  • David James
Yusuf Johari
University of Sheffield

Corresponding Author:[email protected]

Author Profile
Stephen Jaffe
University of Sheffied
Author Profile
Joseph Scarrott
University of Sheffield
Author Profile
Abayomi Johnson
University of Sheffield
Author Profile
Théo Mozzanino
University of Sheffield
Author Profile
Thilo Pohle
University of Sheffield
Author Profile
Sheetal Maisuria
Sheffield Teaching Hospitals NHS Foundation Trust
Author Profile
Amina Bhayat-Cammack
Sheffield Teaching Hospitals NHS Foundation Trust
Author Profile
Adam Brown
University of Sheffield
Author Profile
Kang Lan Tee
University of Sheffield
Author Profile
Philip Jackson
University of Sheffield
Author Profile
Tuck Seng Wong
University of Sheffield
Author Profile
Mark Dickman
University of Sheffield
Author Profile
Ravishankar Sargur
Sheffield Teaching Hospitals NHS Foundation Trust
Author Profile
David James
University of Sheffield
Author Profile

Abstract

We describe scalable and cost-efficient production of full length, His-tagged SARS-CoV-2 spike glycoprotein trimer by CHO cells that can be used to detect SARS-CoV-2 antibodies in patient sera at high specificity and sensitivity. Transient production of spike in both HEK and CHO cells mediated by PEI was increased significantly (up to 10.9-fold) by a reduction in culture temperature to 32ÂșC to permit extended duration cultures. Based on these data GS-CHO pools stably producing spike trimer under the control of a strong synthetic promoter were cultured in hypothermic conditions with combinations of bioactive small molecules to increase yield of purified spike product 4.9-fold to 53 mg/L. Purification of recombinant spike by Ni-chelate affinity chromatography initially yielded a variety of co-eluting protein impurities identified as host cell derived by mass spectrometry, which were separated from spike trimer using a modified imidazole gradient elution. Purified CHO spike trimer antigen was used in ELISA format to detect IgG antibodies against SARS-CoV-2 in sera from patient cohorts previously tested for viral infection by PCR, including those who had displayed COVID-19 symptoms. The antibody assay, validated to ISO 15189 Medical Laboratories standards, exhibited a specificity of 100% and sensitivity of 92.3%. Our data show that CHO cells are a suitable host for the production of larger quantities of recombinant SARS-CoV-2 trimer which can be used as antigen for mass serological testing.
05 Aug 2020Submitted to Biotechnology and Bioengineering
05 Aug 2020Submission Checks Completed
05 Aug 2020Assigned to Editor
03 Sep 2020Reviewer(s) Assigned
25 Sep 2020Review(s) Completed, Editorial Evaluation Pending
25 Sep 2020Editorial Decision: Revise Major
14 Oct 20201st Revision Received
14 Oct 2020Submission Checks Completed
14 Oct 2020Assigned to Editor
22 Oct 2020Reviewer(s) Assigned
27 Oct 2020Review(s) Completed, Editorial Evaluation Pending
27 Oct 2020Editorial Decision: Accept
Feb 2021Published in Biotechnology and Bioengineering volume 118 issue 2 on pages 1013-1021. 10.1002/bit.27615