Production of Trimeric SARS-CoV-2 Spike Protein by CHO Cells for Serological COVID-19 Testing.

Johari, Yusuf B; Jaffé, Stephen R P; Scarrott, Joseph M; Johnson, Abayomi O; Mozzanino, Théo; Pohle, Thilo H; Maisuria, Sheetal; Bhayat-Cammack, Amina; Lambiase, Giulia; Brown, Adam J; Tee, Kang Lan; Jackson, Philip J; Wong, Tuck Seng; Dickman, Mark J; Sargur, Ravishankar; James, David C

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. This article is protected by copyright. All rights reserved.