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Journal of immunological methods

Journal of immunological methods

IF: 1.6
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The spike-ACE2 binding assay: An in vitro platform for evaluating vaccination efficacy and for screening SARS-CoV-2 inhibitors and neutralizing antibodies.

Published:1 April 2022 DOI: 10.1016/j.jim.2022.113244
Shuangzhe Zhang , Chunhui Gao , Tuhin Das , Shuhong Luo , Hao Tang , Xinyi Yao , Chih Yun Cho , Jingqiao Lv , Kino Maravillas , Valerie Jones , Xiaofeng Chen , Ruopan Huang

Abstract

Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has become a worldwide pandemic, and there is a pressing need for the rapid development of novel therapeutic strategies. SARS-CoV-2 viral entry is mediated by interaction between the receptor binding domain (RBD) of the SARS-CoV-2 Spike protein and host cellular receptor, human angiotensin converting enzyme 2 (ACE2). The lack of a high throughput screening (HTS) platform for candidate drug screening means that no targeted COVID-19 treatments have been developed to date. To overcome this limitation, we developed a novel, rapid, simple, and HTS binding assay platform to screen potential inhibitors of the RBD-ACE2 complex. Three “neutralizing” mouse monoclonal antibodies capable of blocking the RBD-ACE2 interaction were identified using our binding assay and pseudovirus neutralization assay followed by further validation with the Focus Reduction Neutralization Test (FRNT), which analyzes the neutralization capacity of samples in the presence of live SARS-CoV-2. Furthermore, the consistency of our binding assay and FRNT results (R2?=?0.68) was demonstrated by patients' serum, of which were COVID-19 positive (n?=?34) and COVID-19 negative (n?=?76). Several small molecules selected for their potential to inhibit the Spike-ACE2 complex in silico were also confirmed with the binding assay. In addition, we have evaluated vaccine efficacy using binding assay platform and validated through pseudovirus neutralization assay. The correlation between binding assay & psuedovirus assay of the post vaccinated serum showed well correlated (R2?=?0.09) Moreover, our binding assay platform successfully validated different Spike RBD mutants. These results indicate that our binding assay can be used as a platform for in vitro screening of small molecules and monoclonal antibodies, and high-throughput assessment of antibody levels after vaccination. When conducting drug screening, computer virtual screening lacks actual basis, construction of pseudoviruses is relatively complicated, and even FRNT requires a P3 laboratory. There are few methods to determine the competitiveness of the target drug and SRBD or ACE2. Our binding assay can fill this gap and accelerate the process and efficiency of COVID-19 drug screening.

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