Luciferase readout: Raw neutralization results for neutralization assays from pseudoparticles containing the SARS-CoV-2 receptor binding domain from a cryptic lineage
Data files
Mar 08, 2024 version files 66.39 KB
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MO45_G._Luc_Neutralizations_Torin_Hunter.xlsx
65.95 KB
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README.md
441 B
Abstract
Deep sequencing of wastewater to detect SARS-CoV-2 has been used during the COVID- 19 pandemic to monitor viral variants as they appear and circulate in communities. SARS- CoV-2 lineages of an unknown source that have not been detected in clinical samples, referred to as cryptic lineages, are sometimes repeatedly detected from specific locations. We have continued to detect one such lineage previously seen in a Missouri site. This cryptic lineage has continued to evolve, indicating continued selective pressure similar to that observed in Omicron lineages.
This file contains the raw neutralization data using pseudoparticles containing a SARS-CoV-2 Spike protein with the RBD from the cryptic lineage detected in Missouri wastewater.
README: Research data
https://doi.org/10.5061/dryad.4tmpg4fj3
File contains the raw luciferase readout from neutralization assays with the pseudoparticles with the cryptic lineage spike protein.
Description of the data and file structure
The first tab contains detailed data description.
Tabs 2-4 are the raw and process output from 3 independent experiments.
Tab 5 is the average of the three.
Methods
During the viral neutralization assays, 96-well plates were prepared for mAb assessment. 50 μL of supplemented DMEM was added to wells 2-12. The first well of each plate received 100 μL of only antibody supernatant, and then the subsequent wells were serially diluted by transfer- ring supernatant from the previous well. At the tenth well of each row, 50 μL of the dilution was discarded, ensuring a consistent volume of 50 μL in all wells.
Next, 50 μL of viral supernatant was added to wells 1-11 so that wells 11 in all rows serve as positive controls containing only virus, media, and cells (added after incubation). In contrast, wells 12 were negative controls with only cells (added after incubation) and media. The plates were incubated at 37 ̊C with 5% CO2 for at least 1 hour. Following incubation, ~28,000 of the 293FT/ACE2/TMPRSS2 cells were added to each well of the 96-well plates and then incubated for 2 days. All neutralizations were performed in triplicate on each plate.
Two days post-infection, 20 μL of supernatant from all wells was collected and placed into a black 96-well plate. Then, 50 μl of 25 mM coelenterazine (CTZ; Nanolight Technology) sus- pended in phosphate-buffered saline (PBS; Cytiva) was added to all wells and placed into a Per- kinElmer EnSpire 2300 Multilabel Reader to measure Gaussia luciferase activity. Infectivity was measured as relative light units (RLU).