A rapid and efficient screening system for neutralizing antibodies and its application for the discovery of potent neutralizing antibodies to SARS-CoV-2 S-RBD

Abstract

Neutralizing antibodies (Abs) have been considered as promising therapeutics for the prevention and treatment of pathogens. After the outbreak of COVID-19, potent neutralizing Abs to SARS-CoV-2 were promptly developed, and a few of those neutralizing Abs are being tested in clinical studies. However, there were few methodologies detailly reported on how to rapidly and efficiently generate neutralizing Abs of interest. Here, we present a strategically optimized method for precisive screening of neutralizing monoclonal antibodies (mAbs), which enabled us to identify SARS-CoV-2 receptor-binding domain (RBD) specific Abs within 4 days, followed by another 2 days for neutralization activity evaluation. By applying the screening system, we obtained 198 Abs against the RBD of SARS-CoV-2. Excitingly, we found that approximately 50% (96/198) of them were candidate neutralizing Abs in a preliminary screening of SARS-CoV-2 pseudovirus and 20 of these 96 neutralizing Abs were confirmed with high potency. Furthermore, 2 mAbs with the highest neutralizing potency were identified to block authentic SARS-CoV-2 with the half-maximal inhibitory concentration (IC50) at concentrations of 9.88 ng/ml and 11.13 ng/ml. In this report, we demonstrated that the optimized neutralizing Abs screening system is useful for the rapid and efficient discovery of potent neutralizing Abs against SARS-CoV-2. Our study provides a methodology for the generation of preventive and therapeutic antibody drugs for emerging infectious diseases.

A key linear epitope for a potent neutralizing antibody to SARS-CoV-2 S-RBD

“…2b, Table 1). Two of them (58G6 and 510A5) showed consistent results with our previous findings 6 . The other 16 potential NAbs blocked authentic SARS-CoV-2 with the IC 50 ranging from 0.043 to 0.153 μg/mL, and they neutralized SARS-CoV-2 pseudovirus with similar IC50 (Extended Data Fig.…”

“…In our previous study, at least 50% S-RBD specific mAbs derived from the memory B cells were found to be NAbs 6 . Here, we focused on the top 20 potential NAbs for their functional characterization and the neutralizing mechanism study, in detail.…”

“…Previously, we screened a total of 219 mAbs to SARS-CoV-2 S-RBD from the memory B cells of COVID-19 convalescent patients via our established NAbs screening system, and discovered a panel of potential NAbs by the authentic virus cytopathic effect (CPE) assay 6 . The top 20 of these potential NAbs were used for functional characterization.…”

“…Due to the fact that clinically available NAbs and vaccines are currently under investigations, their safety and efficacy remain largely unknown. Independently, our group and others have reported that NAbs could efficiently block authentic virus entry by interfering with SARS-CoV-2 S-RBD and its receptor binding 6,15–19 . Several studies have demonstrated the epitope mapping of the NAbs to SARS-CoV-2 in three dimensions, which has been proved to promote the development of effective vaccines for COVID-19 15–18 .…”

“…Its variant carrying mutated Spike (S) protein D614G (S D614G ) has become the most prevalent form in the current global pandemic 4,5 . We have identified a large panel of potential neutralizing antibodies (NAbs) targeting the receptor-binding domain (RBD) of SARS-CoV-2 S 6 . Here, we focused on the top 20 potential NAbs for the mechanism study.…”