Simultaneous evaluation of antibodies that inhibit SARS-CoV-2 variants via multiplex assay
Ester Lopez, … , Wai-Hong Tham, Amy W. Chung
Ester Lopez, … , Wai-Hong Tham, Amy W. Chung
Published July 12, 2021
Citation Information: JCI Insight. 2021;6(16):e150012. https://doi.org/10.1172/jci.insight.150012.
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Resource and Technical Advance COVID-19

Simultaneous evaluation of antibodies that inhibit SARS-CoV-2 variants via multiplex assay

Abstract

The SARS-CoV-2 receptor binding domain (RBD) is both the principal target of neutralizing antibodies and one of the most rapidly evolving domains, which can result in the emergence of immune escape mutations, limiting the effectiveness of vaccines and antibody therapeutics. To facilitate surveillance, we developed a rapid, high-throughput, multiplex assay able to assess the inhibitory response of antibodies to 24 RBD natural variants simultaneously. We demonstrate how this assay can be implemented as a rapid surrogate assay for functional cell-based serological methods to measure the SARS-CoV-2 neutralizing capacity of antibodies at the angiotensin-converting enzyme 2–RBD (ACE2-RBD) interface. We describe the enhanced affinity of RBD variants N439K, S477N, Q493L, S494P, and N501Y to the ACE2 receptor and demonstrate the ability of this assay to bridge a major gap for SARS-CoV-2 research, informing selection of complementary monoclonal antibody candidates and the rapid identification of immune escape to emerging RBD variants following vaccination or natural infection.

Authors

Ester Lopez, Ebene R. Haycroft, Amy Adair, Francesca L. Mordant, Matthew T. O’Neill, Phillip Pymm, Samuel J. Redmond, Wen Shi Lee, Nicholas A. Gherardin, Adam K. Wheatley, Jennifer A. Juno, Kevin J. Selva, Samantha K. Davis, Samantha L. Grimley, Leigh Harty, Damian F.J. Purcell, Kanta Subbarao, Dale I. Godfrey, Stephen J. Kent, Wai-Hong Tham, Amy W. Chung

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Figure 4

Mapping mutations to the RBD that affect recognition by mAbs.

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Mapping mutations to the RBD that affect recognition by mAbs. (A) Relati...
(A) Relative EC50 RBD binding of each mAb to each of the RBD variants. (B) Relative IC50 RBD inhibition of each mAb to each of the RBD variants. (C) RBD natural variant ACE2-RBD relative IC50 inhibition relative to RBD WT. Blue, stronger inhibition of RBD variants relative to WT (IC50 < RBD WT). Orange, weaker inhibition of RBD variants relative to WT (>1- to 6-fold RBD WT IC50). Red, more than 6-fold weaker RBD WT IC50, i.e., very poor/absence of inhibition. (D) Relative IC50 RBD inhibition of each mAb to each of the RBD variants performed in a noncompetitive format where mAbs were preincubated prior to addition of ACE2. (E) Wilcoxon’s paired 2-tailed t test comparison of IC50 values obtained via microneutralization assay for virus preincubated with mAb for 1 hour at 37°C (preincubated) versus addition of mAb, virus, and cells (combined). (F) Relative EC50 binding of mAbs to alanine (or serine if previously alanine) RBD mutants generated of selected RBD variants via multiplex. **P < 0.01.