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. 2021 Feb 10;11:3461. doi: 10.1038/s41598-021-83019-0

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© The Author(s) 2021

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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Anti-SARS-CoV-2-RBD neutralization antibodies using spotted electrochemiluminescence ELISA. Peripheral blood was collected from hospitalized COVID-19 and anonymous recovered patients (n = 75). Negative samples were obtained from true SARS-CoV-2 negative patients (i.e., prior to the SARS-CoV-2 pandemic) (n = 4). Plasma was obtained, diluted 1:50, and added to a 10-spot 96-well plate spotted with SARS-CoV-2 RBD antigen on spot number 1, and BSA on spots number 2–10. ACE2-sulfotag was used instead of a secondary/detection antibody. Representative photomicrographs of real time (a) and schematic (b) images of the spotted 96 well plate are presented. Kinetics of all patients (c), mild (d) and moderate/severe (e) patients is shown; average ± SEM. Inhibition of ACE2-RBD binding was calculated from the average RLU of the 4 negative plasma donors. (f–h) Patients’ antibody results were graphed against the neutralization antibody response. (i–k) Correlation analysis of antibody vs. ACE-RBD binding. Data were calculated using GraphPad Prism 8; the dotted X-line represents the calculated cutoff values (95% and 98% sensitivity) discriminating between positive and negative samples. (c–e) A Nonparametric Kruskal–Wells test for multiple comparisons. (i–k) Correlation analysis was performed using a nonparametric Spearman’s correlation test (two-tailed, 95% confidence). P values are shown.