Monoclonal antibodies binding data for SARS-CoV-2 proteins

Published: 5 May 2022| Version 9 | DOI: 10.17632/9hns9ss6ny.9
Contributors:
Nawneet Mishra,
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Description

ELISA EC50 The raw dataset of EC50 for the selected IgGs that are specific for SARS2-CoV-2 proteins. Data points and analyzed set were included in the raw data in excel format. The data were fit to standard four-parameter logistic equations using GraphPad Prism (GraphPad Software, La Jolla, CA). BLI (Bio-Layer Interferometry) The raw data are in excel format which includes data points and fitting points for selected IgGs that are specific for SARS2-CoV-2 proteins. The binding responses were analyzed by subtracting data from reference, and data were globally fitted with a 1:1 binding model using ForteBio’s Data Analysis software 9.0.

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Steps to reproduce

ELISA EC50 384-well microplates were coated overnight at 4 oC with 2 μg/mL protein or GST/MHT control proteins in PBS pH 7.4. Wells were blocked with 0.2% BSA in PBS for one hour and washed 4 times with 0.05% Tween in PBS (PT buffer). Serial diluted IgGs were into PBT transferred to antigen-coated plates and incubated at room temperature for 30 min, washed with PBST four times, and incubated for 30 min with anti-β-HRP antibody conjugate (1:7500 dilution in PBT). The plate was washed, developed, and read as described above. The data were fit to standard four-parameter logistic equations using GraphPad Prism (GraphPad Software, La Jolla, CA) and EC50 values were estimated. Binding Kinetics (BLI) BLI experiments were performed on an Octet HTX instrument (Sartorius) at 1000 rpm and 25 °C, were 2-20 microgram per milliliter of SARS-CoV2 antigens were covalently captured on AR2G biosensors (Sartorius, 18-5092) by amine-reactive coupling chemistry, and unbound reactive N-hydroxysulfosuccinimide (NHS) esters on the biosensors were quenched for 600 s by 1M ethanolamine (pH 8.5)[3, 4]. The biosensor probe were equilibrating with assay buffer (PBS, 1% BSA, 0.05% Tween 20), dipped for 600 s into wells containing serial 3-fold dilutions of IgGs, and were transferred back into assay buffer for 600 s of dissociation. The binding responses were analyzed by subtracting data from reference, and data were globally fitted with 1:1 binding model using ForteBio’s Data Analysis software 9.0.

Institutions

Terrence Donnelly Centre for Cellular and Biomolecular Research, Washington University in Saint Louis, Boston University School of Medicine

Categories

Biochemical Analysis, Western Blot

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