Extracts were prepared according to published protocols using sonication and postlysis processing in the Rosetta2 (DE3) pLysS strain10 . Briefly, cells are plated on a chloramphenicol-selective agar plate and incubated overnight then used to inoculate a 20 mL overnight starter culture for a 1 L final culture. This culture is grown to an optical density (OD600) of 3.0 ± 0.2 then pelleted and lysed by sonication before centrifugation for 10 minutes at 4°C and 12,000g. After lysis, extracts were incubated with shaking for 80 minutes at 37°C and 200 rpm then recentrifuged under the same conditions. The supernatant was injected into a 10K MWCO dialysis cassette (ThermoFisher, 66380) and dialyzed at 4°C for three hours before a final centrifugation under the same conditions and snap-freezing in liquid nitrogen.
10k mwco dialysis cassette
Manufactured by Thermo Fisher Scientific
Sourced in United States
The 10K MWCO dialysis cassette is a laboratory equipment used for dialysis, a process that removes small molecules and ions from a solution while retaining larger molecules. The cassette has a molecular weight cut-off (MWCO) of 10,000 Daltons, which means it effectively retains molecules larger than 10,000 Daltons while allowing smaller molecules to pass through. This product can be used in various research and purification applications that require the separation and concentration of macromolecules.
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2 protocols using 10k mwco dialysis cassette
1
Recombinant Protein Expression and Purification
2
SARS-CoV-2 RBD Protein Labeling and Detection
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Streptavidin (SA) probe (Gator Bio, CA, USA 160002), 96-well black plates (Greiner Bio-One, Germany 655,209), 96-well Max plates (Gator Bio 130,018), SARS-CoV-2 receptor binding domain protein (RBD, Fc Tag) (SinoBiological, China 40,592-V02H), Albumin Human (Sigma Aldrich, MO, USA A9731), EZ-Link NHS-LC-LC-Biotin (Thermo Fisher Scientific, MA, USA 21343), 10K MWCO Dialysis Cassette (Thermo Fisher Scientific 87,730). PD-10 column (GE Healthcare, IL, USA 17-0851-01). Q Buffer consists of 0.2% BSA, 0.02% Tween-20, 0.05% Sodium azide in PBS pH 7.4, Gator Bio 120,010. Rabbit SARS-CoV-2 Spike antibody (Sinobiological 40,591-T62).
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