Optimizing high-yield production of SARS-CoV-2 soluble spike trimers for serology assays
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SciScore for 10.1101/2020.05.27.120204: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement not detected. Randomization not detected. Blinding not detected. Power Analysis not detected. Sex as a biological variable not detected. Table 2: Resources
No key resources detected.
Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).
Results from LimitationRecognizer: We detected the following sentences addressing limitations in the study:Previous work in our lab and others suggests one limitation to protein secretion in transiently transfected mammalian cell culture might be the secretion process itself. While multiple factors appear to be involved, a recent …
SciScore for 10.1101/2020.05.27.120204: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement not detected. Randomization not detected. Blinding not detected. Power Analysis not detected. Sex as a biological variable not detected. Table 2: Resources
No key resources detected.
Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).
Results from LimitationRecognizer: We detected the following sentences addressing limitations in the study:Previous work in our lab and others suggests one limitation to protein secretion in transiently transfected mammalian cell culture might be the secretion process itself. While multiple factors appear to be involved, a recent paper suggests that limiting components in the secretory pathway appear to be the major bottleneck [9,10]. We did observe a significant amount of spike protein in the Expi293 cell pellets (data not shown), suggesting that some protein was being held up in the endoplasmic reticulum, or was otherwise failing to completely mature through the secretory pathway. Low cell viabilities (<70%) at harvest were another sign of potential toxicity caused by secretory failures. Thus, we compared the purification yield of VRC and Mt. Sinai spike from transiently transfected cells incubated at either 32°C or 37°C after the addition of enhancers 18-hours post-transfection. As seen in Table 1 and Fig. 2B, the lower temperature expression led to a dramatic increase in yield to ∼5 mg/l for both CoV-2 spike proteins we tested. This yield is similar to that cited in a recent report of the Mt. Sinai spike protein produced at 37°C and using a diafiltration approach to buffer exchange [3]. However, unpublished results from other colleagues in the field using this construct suggest 1-2 mg/l is a more consistently observed result, suggesting that these improvements will make a marked enhancement to yield. Published VRC spike protein yields are 0.5 mg/l [2], suggesting that our modi...
Results from TrialIdentifier: No clinical trial numbers were referenced.
Results from Barzooka: We did not find any issues relating to the usage of bar graphs.
Results from JetFighter: We did not find any issues relating to colormaps.
Results from rtransparent:- Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
- Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
- No protocol registration statement was detected.
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