Effect of Inactivation Methods on SARS-CoV-2 Virion Protein and Structure
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Abstract
The risk posed by Severe Acute Respiratory Syndrome Coronavirus -2 (SARS-CoV-2) dictates that live-virus research is conducted in a biosafety level 3 (BSL3) facility. Working with SARS-CoV-2 at lower biosafety levels can expedite research yet requires the virus to be fully inactivated. In this study, we validated and compared two protocols for inactivating SARS-CoV-2: heat treatment and ultraviolet irradiation. The two methods were optimized to render the virus completely incapable of infection while limiting the destructive effects of inactivation. We observed that 15 min of incubation at 65 °C completely inactivates high titer viral stocks. Complete inactivation was also achieved with minimal amounts of UV power (70,000 µJ/cm2), which is 100-fold less power than comparable studies. Once validated, the two methods were then compared for viral RNA quantification, virion purification, and antibody detection assays. We observed that UV irradiation resulted in a 2-log reduction of detectable genomes compared to heat inactivation. Protein yield following virion enrichment was equivalent for all inactivation conditions, but the quality of resulting viral proteins and virions were differentially impacted depending on inactivation method and time. Here, we outline the strengths and weaknesses of each method so that investigators might choose the one which best meets their research goals.
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SciScore for 10.1101/2020.11.14.383026: (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. Cell Line Authentication not detected. Table 2: Resources
Antibodies Sentences Resources Primary antibody complexes were detected with goat anti-mouse (Invitrogen) or goat anti-rabbit (Santa Cruz Biotech) HRP conjugated secondary antibodies. anti-mousesuggested: Noneanti-rabbitsuggested: NoneAntisera was decanted and plates were washed as above, followed by incubation for 1 hour at room temperature with a 1:3000 dilution of goat-anti-rabbit secondary antibody conjugated to HRP in wash buffer with 1% nonfat milk. secondarysuggested: None… SciScore for 10.1101/2020.11.14.383026: (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. Cell Line Authentication not detected. Table 2: Resources
Antibodies Sentences Resources Primary antibody complexes were detected with goat anti-mouse (Invitrogen) or goat anti-rabbit (Santa Cruz Biotech) HRP conjugated secondary antibodies. anti-mousesuggested: Noneanti-rabbitsuggested: NoneAntisera was decanted and plates were washed as above, followed by incubation for 1 hour at room temperature with a 1:3000 dilution of goat-anti-rabbit secondary antibody conjugated to HRP in wash buffer with 1% nonfat milk. secondarysuggested: NoneExperimental Models: Cell Lines Sentences Resources Viral stocks were propagated and titered on E6 Vero cells in DMEM supplemented with 2% FBS and 1% pen-strep. Verosuggested: CLS Cat# 605372/p622_VERO, RRID:CVCL_0059)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: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.Results from TrialIdentifier: No clinical trial numbers were referenced.
Results from Barzooka: We found bar graphs of continuous data. We recommend replacing bar graphs with more informative graphics, as many different datasets can lead to the same bar graph. The actual data may suggest different conclusions from the summary statistics. For more information, please see Weissgerber et al (2015).
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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