A novel competition ELISA for the rapid quantification of SARS‐CoV‐2 neutralizing antibodies in convalescent plasma

Abstract

Background: COVID-19 convalescent plasma (CCP) ideally contains high titers of (neutralizing) anti-SARS-CoV-2 antibodies. Several scalable immunoassays for CCP selection have been developed. We designed an enzyme-linked immunosorbent assay (ELISA) that measures neutralizing antibodies (of all isotypes) in plasma by determining the level of competition between CCP and a mouse neutralizing antibody for binding to the receptor binding domain (RBD) of SARS-CoV-2.Methods: Plasma was collected from 72 convalescent individuals and inhibition of viral infection was determined by plaque reduction neutralization (PRNT50). The level of neutralizing antibodies was measured in the novel competition ELISA and in a commercially available ELISA that measures inhibition of recombinant ACE2 binding to immobilized RBD. These results were compared with a high throughput chemiluminescent microparticle immunoassay (CMIA). Results: The results from both ELISAs were correlating, in particular for high titer CCP (PRNT50 ≥ 1:160) (Spearman r = .73, p < .001). Moderate correlation was found between the competition ELISA and CMIA (r = .57 for high titer and r = .62 for low titer CCP, p < .001). Receiver operator characteristic analysis showed that the competition ELISA selected CCP with a sensitivity and specificity of 61% and 100%, respectively. However, discrimination between low and high titer CCP had a lower resolution (sensitivity: 34% and specificity: 89%). Conclusion:The competition ELISA screens for neutralizing antibodies in CCP by competition for just a single epitope. It exerts a sensitivity of 61% with

A Serological Multiplexed Immunoassay (MIA) Detects Antibody Reactivity to SARS-CoV-2 and Other Viral Pathogens in Liberia and Is Configurable as a Multiplexed Inhibition Test (MINT)

“…Several ELISA-based SARS-CoV-2 inhibition assays have been developed to determine surrogate neutralization without the need for BSL-3 laboratories [15,[37][38][39]. We developed an inhibition assay to complement our MIA panel and capitalize on the multiplexed functionality of bead-based systems.…”

Latex Microsphere-Based Bicolor Immunochromatography for Qualitative Detection of Neutralizing Antibody against SARS-CoV-2

“…Live virus- or pseudovirus-based neutralization testing assays are not implemented in routine practice. Recently, ELISA based Nab that interacts with ACE2 protein in vitro has been established [ 24 ], but the operation of ELISA is tedious, time-consuming, and depends on special equipment. Finding alternative methods without cell and virus culture operation is of interest to obtain reliable NAb information for in vitro assays, which are simple, fast, high-throughput, and commercially available.…”

“…However, it is highly technical, often time consuming, and high risk, because live virus- or pseudovirus-based neutralization testing is restricted to biosafety level 3 and 2 facilities. Although an enzyme-linked immunosorbent assay (ELISA) has also been developed for accurate and sensitive detection of SARS-CoV-2 antibodies, its labor- and time-consuming operation limits its widespread application on site [ 22 , 23 , 24 ]. In practice, a direct and rapid detection method without pretreatment is deserved for point-of-care testing of NAb among vaccine recipients.…”

Intranasal administration of convalescent plasma protects against SARS-CoV-2 infection in hamsters

“…The PRNT was performed as previously described. 10 , 22 Variants used were SARS-CoV-2 ancestral strain (BetaCov/Belgium/GHB-03021/202, EPI ISL 407976|2020-02-03, passage 5), 23 Delta B.1.617.2 (hCoV-19/Belgium/rega-7214/2021; EPI_ISL_2425097, passage 2) 24 , 25 and Omicron B.1.1.529 (hCoV-19/Belgium/rega-20174/2021, EPI_ISL_6794907|2021, passage 2). 26 In brief, a ten-step serial dilution was prepared for samples CCP, VCCP and NIP HUMAN .…”