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Production of Trimeric SARS-CoV-2 Spike Protein by CHO Cells for Serological COVID-19 Testing
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  • Yusuf Johari,
  • Stephen Jaffe,
  • Joseph Scarrott,
  • Abayomi Johnson,
  • Théo 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]

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Stephen Jaffe
University of Sheffied
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Joseph Scarrott
University of Sheffield
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Abayomi Johnson
University of Sheffield
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Théo Mozzanino
University of Sheffield
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Thilo Pohle
University of Sheffield
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Sheetal Maisuria
Sheffield Teaching Hospitals NHS Foundation Trust
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Amina Bhayat-Cammack
Sheffield Teaching Hospitals NHS Foundation Trust
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Adam Brown
University of Sheffield
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Kang Lan Tee
University of Sheffield
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Philip Jackson
University of Sheffield
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Tuck Seng Wong
University of Sheffield
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Mark Dickman
University of Sheffield
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Ravishankar Sargur
Sheffield Teaching Hospitals NHS Foundation Trust
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David James
University of Sheffield
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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 32ºC 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