Optimizing high-yield production of SARS-CoV-2 soluble spike trimers for serology assays
The SARS-CoV-2 spike trimer is the primary antigen for several serology assays critical to determining the extent of SARS-CoV-2 exposure in the population. Until stable cell lines are developed to increase the titer of this secreted protein in mammalian cell culture, the low yield of spike protein produced from transient transfection of HEK293 cells will be a limiting factor for these assays. To improve the yield of spike protein and support the high demand for antigens in serology assays, we investigated several recombinant protein expression variables by altering the incubation temperature, harvest time, chromatography strategy, and final protein manipulation. Through this investigation, we developed a simplified and robust purification strategy that consistently yields 5 mg of protein per liter of expression culture for two commonly used forms of the SARS-CoV-2 spike protein. We show that these proteins form well-behaved stable trimers and are consistently functional in serology assays across multiple protein production lots.
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UpdateIn: Protein Expr Purif. 2020 Oct;174:105686. - PMID 32504802 |
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| Medienart: |
E-Artikel |
| Erscheinungsjahr: |
2020 |
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| Erschienen: |
2020 |
| Enthalten in: |
Zur Gesamtaufnahme - year:2020 |
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| Enthalten in: |
bioRxiv : the preprint server for biology - (2020) vom: 28. Mai |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Esposito, Dominic [VerfasserIn] |
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| Links: |
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| Themen: |
COVID-19 |
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Date Revised 19.10.2023 published: Electronic UpdateIn: Protein Expr Purif. 2020 Oct;174:105686. - PMID 32504802 Citation Status PubMed-not-MEDLINE |
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| doi: |
10.1101/2020.05.27.120204 |
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| PPN (Katalog-ID): |
NLM310908884 |
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| 500 | |a UpdateIn: Protein Expr Purif. 2020 Oct;174:105686. - PMID 32504802 | ||
| 500 | |a Citation Status PubMed-not-MEDLINE | ||
| 520 | |a The SARS-CoV-2 spike trimer is the primary antigen for several serology assays critical to determining the extent of SARS-CoV-2 exposure in the population. Until stable cell lines are developed to increase the titer of this secreted protein in mammalian cell culture, the low yield of spike protein produced from transient transfection of HEK293 cells will be a limiting factor for these assays. To improve the yield of spike protein and support the high demand for antigens in serology assays, we investigated several recombinant protein expression variables by altering the incubation temperature, harvest time, chromatography strategy, and final protein manipulation. Through this investigation, we developed a simplified and robust purification strategy that consistently yields 5 mg of protein per liter of expression culture for two commonly used forms of the SARS-CoV-2 spike protein. We show that these proteins form well-behaved stable trimers and are consistently functional in serology assays across multiple protein production lots | ||
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| 700 | 1 | |a Mehalko, Jennifer |e verfasserin |4 aut | |
| 700 | 1 | |a Drew, Matthew |e verfasserin |4 aut | |
| 700 | 1 | |a Snead, Kelly |e verfasserin |4 aut | |
| 700 | 1 | |a Wall, Vanessa |e verfasserin |4 aut | |
| 700 | 1 | |a Taylor, Troy |e verfasserin |4 aut | |
| 700 | 1 | |a Frank, Peter |e verfasserin |4 aut | |
| 700 | 1 | |a Denson, John-Paul |e verfasserin |4 aut | |
| 700 | 1 | |a Hong, Min |e verfasserin |4 aut | |
| 700 | 1 | |a Gulten, Gulcin |e verfasserin |4 aut | |
| 700 | 1 | |a Sadtler, Kaitlyn |e verfasserin |4 aut | |
| 700 | 1 | |a Messing, Simon |e verfasserin |4 aut | |
| 700 | 1 | |a Gillette, William |e verfasserin |4 aut | |
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