SARS-CoV-2 RBD and Ovalbumin Biotinylation

Purified and Avi-tagged SARS-CoV-2 Wuhan-Hu-1 RBD was biotinylated using the Biotin-Protein Ligase-BIRA kit according to manufacturer’s instructions (Avidity) as described before^{16 (link)}. Ovalbumin (Sigma, A5503-1G) was biotinylated using the EZ-Link Sulfo-NHS-LC-Biotinylation kit according to the manufacturer’s instructions (Thermo Scientific). Biotinylated ovalbumin was conjugated to streptavidin-BV711 for single-cell sorts (BD biosciences, 563262) or to streptavidin-BB515 for phenotyping panel (BD biosciences, 564453). RBD was conjugated to streptavidin-PE (BD Biosciences, 554061) and streptavidin-AF647 (Biolegend, 405237)^{16 (link)}.

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Muecksch F., Wang Z., Cho A., Gaebler C., Ben Tanfous T., DaSilva J., Bednarski E., Ramos V., Zong S., Johnson B., Raspe R., Schaefer-Babajew D., Shimeliovich I., Daga M., Yao K.H., Schmidt F., Millard K.G., Turroja M., Jankovic M., Oliveira T.Y., Gazumyan A., Caskey M., Hatziioannou T., Bieniasz P.D, & Nussenzweig M.C. (2022). Increased memory B cell potency and breadth after a SARS-CoV-2 mRNA boost. Nature, 607(7917), 128-134.

Publication 2022

Biotin-Protein Ligase-BIRA kit EZ-Link Sulfo-NHS-LC-Biotinylation kit streptavidin-BV711 streptavidin-BB515 streptavidin-PE streptavidin-AF647

Af647 Biotin ligase Biotinylation Cell Ovalbumin Protein Sars cov 2 Streptavidin Sulfo nhs

Corresponding Organization :

Other organizations : Rockefeller University, Howard Hughes Medical Institute

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Variable analysis

independent variables

Biotinylation of purified and Avi-tagged SARS-CoV-2 Wuhan-Hu-1 RBD using the Biotin-Protein Ligase-BIRA kit
Biotinylation of ovalbumin using the EZ-Link Sulfo-NHS-LC-Biotinylation kit

dependent variables

Conjugation of biotinylated ovalbumin to streptavidin-BV711 for single-cell sorts
Conjugation of biotinylated ovalbumin to streptavidin-BB515 for phenotyping panel
Conjugation of RBD to streptavidin-PE
Conjugation of RBD to streptavidin-AF647

control variables

Manufacturer's instructions were followed for biotinylation and conjugation procedures

positive controls

Biotinylated ovalbumin conjugated to streptavidin-BV711 and streptavidin-BB515

negative controls

None explicitly mentioned

Annotations

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