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Authors: Chee Wah Tan et al. 2020

Link to paper: https://www.researchsquare.com/article/rs-24574/v1

Journal/ Pre-Print: Nature Research

Tags: COVID-19, serological test, detection of neutralizing antibodies, SARS-CoV-2 surrogate virus neutralization test

Research Highlights 

1. The surrogate neutralisation test is capable of measuring majority neutralising Abs against SARS2 RBD regardless of the isotype - IgG, IgM, IgA - unlike standard ELISAs reported by other groups.

2. Correlation study between infectious virus-based virus neutralization test (VNT) and the RBD-hACE2-based sVNT shows that they are positively correlated.

3. sVNT has been tested against related corona strains (SARS1, MERS, 229/NL63 and OC43) and a slight cross-reaction was detected against serum from SARS-CoV patients. However, the neutralisation capacity of SARS-CoV-2 sera was statistically significantly higher than SARS-CoV.

Summary 

This paper reports on a SARS-CoV-2 surrogate virus neutralization test (sVNT), which assays for total neutralizing antibodies in an isotype- and species-independent manner, via a competition ELISA which detects abs specific to the Spike protein RBD region that block binding to ACE2. sVNT provides a quick (2h) BSL2 assay with a high specificity (100%) and sensitivity of up to 95-100% (depending on the cut-off stringency). This assay has been validated against sera from different corona viruses including SARS1, MERS, 229/NL63 and OC43. Given that most available diagnostic tests are isotype specific, this test has the advantage of measuring the total amount of neutralizing activity in SARS-CoV-2 sera.

Impact for SARS-CoV2/COVID19 research efforts

Develop diagnostic tools for SARS-CoV-2/COVID19: A competition ELISA using ACE2 as a substrate, SARS-CoV-2 S protein RBD conjugated to HRP, and convalescent serum from CoVid-19 patients.

Study Type

· In vitro study

Strengths and limitations of the paper

Novelty: A novel and quick diagnostic test that can be performed at the BSL2 to test neutralisation capacity of SARS-CoV-2 S protein RBD specific antibodies in patient serum.

Standing in the field: Joins other ELISA studies summarised nicely by Infantino et al. here https://www.ima.org.il/FilesUploadPublic/IMAJ/0/423/211570.pdf

Appropriate statistics: ~125 patients or controls in two cohorts. Kruskal-Wallis test/Dunn’s multiple comparisons test. There are few descriptive statistics in the paper and outputs are limited to figures.

Viral model used: Different hCOV viruses were used including SARS1 and MERS. Also 229, NL63 or OC. Different species sera were also tested – rabbits and ferrets.

Translatability: Laboratory based ELISA (BSL2).

Main limitations: Immunogenic epitopes are not only found in RBD but other regions of the S protein and also N protein. This test will only determine those antibodies against epitopes found in the RBD of the S protein of SARS-CoV-2 which block binding to ACE2, thus missing other potentially beneficial effects of antibody binding (e.g. opsonization) or indirect effects of binding described by other groups.

Assay does not discriminate between antibody classes and therefore cannot inform stage of infection (ie. IgM = acute infection, IgG = class switched later in infection) or whether the response is high avidity/low affinity or low avidity/high affinity.

Patient clinical details sparse and timepoints are broadly grouped, common limitations in the current situation.

No information on the quality of the recombinant proteins they are using (all proteins are commercially sourced reagents).