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.
Highlights
SARS-CoV-2 was first identified in late 2019 in Wuhan, China [1, 2], and has since become an ongoing global public health emergency
We used a bead-based multiplex assay to evaluate the neutralizing capacity of SARS-CoV-2 antibodies against recombinant receptor-binding domain (RBD) variants
Convalescent plasma or purified monoclonal antibody (mAb) can be allowed to simultaneously compete with soluble biotin-angiotensin-converting enzyme 2 (ACE2) for binding to the RBD variants coupled to the magnetic beads (Figure 1A)
Summary
SARS-CoV-2 was first identified in late 2019 in Wuhan, China [1, 2], and has since become an ongoing global public health emergency. Binding to ACE2 and entry to the host cell are facilitated via the spike, a homotrimeric transmembrane envelope glycoprotein, which consists of the binding (S1), and transmembrane fusion (S2), domains that are processed from the polyprotein precursor at a polybasic furin cleavage site [9, 12]. Key to this virus–host interaction is the receptor-binding domain (RBD), which lies within the S1 subunit and is critical for binding to ACE2 receptors on target cells [11]. The RBD of SARS-CoV-2 represents a key target for the development of vaccine-elicited humoral immunity, and eliciting potent NAbs capable of blocking ACE2 binding remains a key feature of the development of effective vaccines and antibody therapies
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