A rapid bead-based assay for screening of SARS-CoV-2 neutralizing antibodies
Santhik Subhasingh Lupitha, Pramod Darvin, Aneesh Chandrasekharan, Shankara Narayanan Varadarajan, Soumya Jaya Divakaran, Easwaran Sreekumar, Shijulal Nelson‐Sathi, Perunthottathu K. Umasankar, Sara R. Jones, Iype Joseph, Madhavan Radhakrishna Pillai, T.R. Santhoshkumar
Abstract
Quantitative determination of neutralizing antibodies against Severe Acute Respiratory Syndrome Corona Virus-2 (SARS-CoV-2) is paramount in immunodiagnostics, vaccine efficacy testing, and immune response profiling among the vaccinated population. Cost-effective, rapid, easy-to-perform assays are essential to support the vaccine development process and immunosurveillance studies. We describe a bead-based screening assay for S1-neutralization using recombinant fluorescent proteins of hACE2 and SARS-CoV2-S1, immobilized on solid beads employing nanobodies/metal-affinity tags. Nanobody-mediated capture of SARS-CoV-2-Spike (S1) on agarose beads served as the trap for soluble recombinant ACE2-GFPSpark, inhibited by neutralizing antibody. The first approach demonstrates single-color fluorescent imaging of ACE2-GFPSpark binding to His-tagged S1-Receptor Binding Domain (RBD-His) immobilized beads. The second approach is dual-color imaging of soluble ACE2-GFPSpark to S1-Orange Fluorescent Protein (S1-OFPSpark) beads. Both methods showed a good correlation with the gold standard pseudovirion assay and can be adapted to any fluorescent platforms for screening.