Selection, identification, and characterization of SARS-CoV-2 monoclonal antibody resistant mutants

•We have developed an experimental approach for the selection, identification, and characterization of SARS-CoV-2 MARMs.•This assay can be used to identify AA residues in the S protein required for mNAb-mediated inhibition of viral infection.•Our assay can be used to predict potential natural drift variants that could emerge upon implementation of therapeutic mNAbs.•We also describe experimental approaches to evaluate MARM viral fitness and how to compare them to WT SARS-CoV-2. The use of monoclonal neutralizing antibodies (mNAbs) is being actively pursued as a viable intervention for the treatment of Severe Acute Respiratory Syndrome CoV-2 (SARS-CoV-2) infection and associated coronavirus disease 2019 (COVID-19). While highly potent mNAbs have great therapeutic potential, the ability of the virus to mutate and escape recognition and neutralization of mNAbs represents a potential problem in their use for the therapeutic management of SARS-CoV-2. Studies investigating natural or mNAb-induced antigenic variability in the receptor binding domain (RBD) of SARS-CoV-2 Spike (S) glycoprotein, and their effects on viral fitness are still rudimentary. In this manuscript we described experimental approaches for the selection, identification, and characterization of SARS-CoV-2 monoclonal antibody resistant mutants (MARMs) in cultured cells. The ability to study SARS-CoV-2 antigenic drift under selective immune pressure by mNAbs is important for the optimal implementation of mNAbs for the therapeutic management of COVID-19. This will help to identify essential amino acid residues in the viral S glycoprotein required for mNAb-mediated inhibition of viral infection, to predict potential natural drift variants that could emerge upon implementation of therapeutic mNAbs, as well as vaccine prophylactic treatments for SARS-CoV-2 infection. Additionally, it will also enable the assessment of MARM viral fitness and its potential to induce severe infection and associated COVID-19 disease.