Production and Purification of SARS-CoV-2 RBD

SARS-CoV-2 spike receptor binding domain protein (RBD) was produced by transient transfection of HEK-293F cells with plasmid pCAGGS (32 (link), 33 (link)), containing the SARS-CoV-2 Wuhan-Hu-1 spike glycoprotein gene RBD with C-terminal hexahistidine tag (generously provided by F. Krammer). Transfection was carried utilizing the Expi293 expression system (Gibco, Thermo Fisher Scientific) following the manufacturer’s instructions. RBD was purified by affinity chromatography utilizing nickel-nitrilotriacetic acid (Ni-NTA) agarose (Thermo Fisher Scientific), and RBD purity was assessed by SDS-PAGE.

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Gunawardana M., Breslin J., Cortez JM J.r., Rivera S., Webster S., Ibarrondo F.J., Yang O.O., Pyles R.B., Ramirez C.M., Adler A.P., Anton P.A, & Baum M.M. (2021). Longitudinal COVID-19 Surveillance and Characterization in the Workplace with Public Health and Diagnostic Endpoints. mSphere, 6(4), e00542-21.

Publication 2021

Expi293 expression system nickel-nitrilotriacetic acid ( Ni-NTA) agarose

Agarose chromatography Binding protein Cells Gene protein Hexahistidine Nickel nitrilotriacetic acid Plasmid Sars cov 2 spike glycoprotein Sds page Transfection Transient

Corresponding Organization : Oak Crest Institute of Science

Other organizations : University of California, Los Angeles, The University of Texas Medical Branch at Galveston

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Variable analysis

independent variables

Transfection of HEK-293F cells with plasmid pCAGGS containing the SARS-CoV-2 Wuhan-Hu-1 spike glycoprotein gene RBD with C-terminal hexahistidine tag

dependent variables

Purification of RBD by affinity chromatography utilizing nickel-nitrilotriacetic acid (Ni-NTA) agarose
Assessment of RBD purity by SDS-PAGE

control variables

Expi293 expression system (Gibco, Thermo Fisher Scientific) used for transfection, following the manufacturer's instructions

controls

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