Citation

  • Authors: Tanaka S. et al.
  • Year: 2022
  • Journal: Cell Rep
  • Applications: in vitro / DNA / FectoPRO
  • Cell type: CHO-S
    Description: Chinese hamster ovary cells

Method

FectoPRO was used for suspension CHO-S transfection cultured in CD-CHO medium. CHO-S cells were seeded at a density of 1 x 10^6 cells/mL in 45 mL culture flask. 75 µL of FectoPRO® was mixed with 5 mL of 15 µg/mL plasmid DNA. DNA/FectoPRO® complexes were added to 45 mL of CHO-S culture.

Abstract

The increasing prevalence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants with the ability to escape existing humoral protection conferred by previous infection and/or immunization necessitates the discovery of broadly reactive neutralizing antibodies (nAbs). Utilizing mRNA display, we identify a set of antibodies against SARS-CoV-2 spike (S) proteins and characterize the structures of nAbs that recognize epitopes in the S1 subunit of the S glycoprotein. These structural studies reveal distinct binding modes for several antibodies, including the targeting of rare cryptic epitopes in the receptor-binding domain (RBD) of S that interact with angiotensin-converting enzyme 2 (ACE2) to initiate infection, as well as the S1 subdomain 1. Further, we engineer a potent ACE2-blocking nAb to sustain binding to S RBD with the E484K and L452R substitutions found in multiple SARS-CoV-2 variants. We demonstrate that mRNA display is an approach for the rapid identification of nAbs that can be used in combination to combat emerging SARS-CoV-2 variants.

Pubmed