Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a highly pathogenic zoonotic virus that spreads rapidly. In this work, we improve the hitherto existing neutralization assay system to assess SARS-CoV-2 inhibitors using a pseudo-typed lentivirus coated with the SARS-CoV-2 spike protein (LpVspike +) and angiotensin-converting enzyme 2 (ACE2)-transfected cat Crandell–Rees feline kidney (CRFK) cells as the host cell line. Our method was 10-fold more sensitive compared to the typical human embryonic kidney 293T (HEK293T) cell system, and it was successfully applied to quantify the titers of convalescent antisera and monoclonal anti-spike antibodies required for pseudo virus neutralization. The 50% inhibition dilution (ID50) of two human convalescent sera, SARS-CoV-2 immunoglobulin G (IgG) and SARS-CoV-2 immunoglobulin M (IgM), which were 1:350 (±1:20) and 1:1250 (±1:350), respectively. The 50% inhibitory concentration (IC50) of the IgG, IgM and immunoglobulin A (IgA) anti-SARS-CoV-2 monoclonal antibodies (mAbs) against LpVspike(+) were 0.45 (±0.1), 0.002 (±0.001) and 0.004 (±0.001) µg mL −1 , respectively. We also found that reagents typically used to enhance infection were not effective in the CFRK system. This methodology is both efficient and safe; it can be employed by researchers to evaluate neutralizing monoclonal antibodies and contribute to the discovery of new antiviral inhibitors against SARS-CoV-2.
【저자키워드】 COVID-19, SARS-CoV-2, HIV, ACE2, spike, monoclonal antibody, Neutralization assay, CRFK Cell, pseudo-typed lentivirus, 【초록키워드】 IgG, IgM, coronavirus, Infection, virus, anti-SARS-CoV-2, angiotensin-converting enzyme 2, IC50, Spike protein, kidney, Spread, Immunoglobulin, IgA, lentivirus, zoonotic, Virus neutralization, methodology, convalescent, inhibitor, immunoglobulin A, mAbs, Antisera, neutralizing monoclonal antibody, anti-spike antibody, monoclonal, Dilution, host cell, acute respiratory syndrome, embryonic kidney, inhibitory concentration, 293T, HEK293T, reagent, researcher, human convalescent sera, highly pathogenic, CrFK, effective, Cell, ENhance, IMPROVE, evaluate, required, applied, contribute, antiviral inhibitor, coated with, the SARS-CoV-2, 【제목키워드】 SARS CoV-2, CrFK,