The antibody variable regions, including the heavy and light chain pairing genes, were synthesized by Integrated DNA Technologies. Recombinant antibodies were generated using established protocols as previously described (25 (link)). Antibodies were purified using protein A spin column (CosmoBio), and their concentrations were determined by ELISA using hIgG standard curve.
Protein a spin column
Manufactured by Cosmo Bio
The Protein A spin column is a device used for the purification of antibodies or other proteins that bind to Protein A. It consists of a column containing a Protein A-based resin that selectively binds to the Fc region of immunoglobulins. The column can be centrifuged to separate the bound antibodies from the unbound proteins in the sample.
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Lab products found in correlation
2 protocols using protein a spin column
1
Generation of Recombinant Antibodies
2
Antibody-mediated ACE2 Binding Enhancement
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The SARS-CoV-2 S-NTD binders were expressed and purified using protein A spin column (Cosmo Bio) and concentration was measured using ELISA. ACE2 binding enhancement to Spike WT, Delta, and Omicron variants in the presence of infection enhancing-like antibodies was measured. Briefly, Expi293F cells (Thermo) that express either Spike WT, Delta, or Omicron variant were incubated by 1 µg/mL antibodies for 30 minutes. Followed by ACE2-biotin at 1 µg/mL (RnD Systems) incubation for 30 minutes and then SA-APC (Biolegend) incubation for 1 hour. The amount of ACE2 that binds to Spike protein was measured using flow cytometry (Attune NxT, Thermo). Fold change was calculated by comparing the amount of bound ACE2 in the presence of antibodies and in the absence of antibodies.
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