Abstract
A serological COVID-19 Multiplex Assay was developed and validated using serum samples from convalescent patients and those collected prior to the 2020 pandemic. After initial testing of multiple potential antigens, the SARS-CoV-2 nucleocapsid protein (NP) and receptor-binding domain (RBD) of the spike protein were selected for the human COVID-19 Multiplex Assay. A comparison of synthesized and mammalian expressed RBD proteins revealed clear advantages of mammalian expression. Antibodies directed against NP strongly correlated with SARS-CoV-2 virus neutralization assay titers (rsp = 0.726), while anti-RBD correlation was moderate (rsp = 0.436). Pan-Ig, IgG, IgA, and IgM against NP and RBD antigens were evaluated on the validation sample sets. Detection of NP and RBD specific IgG and IgA had outstanding performance (AUC > 0.90) for distinguishing patients from controls, but the dynamic range of the IgG assay was substantially greater. The COVID-19 Multiplex Assay was utilized to identify seroprevalence to SARS-CoV-2 in people living in a low-incidence community in Ithaca, NY. Samples were taken from a cohort of healthy volunteers (n = 332) in early June 2020. Only two volunteers had a positive result on a COVID-19 PCR test performed prior to serum sampling. Serological testing revealed an exposure rate of at least 1.2% (NP) or as high as 5.7% (RBD), higher than the measured incidence rate of 0.16% in the county at that time. This highly sensitive and quantitative assay can be used for monitoring community exposure rates and duration of immune response following both infection and vaccination.
Publication types
-
Research Support, Non-U.S. Gov't
MeSH terms
-
Adolescent
-
Adult
-
Aged
-
Aged, 80 and over
-
Antibodies, Neutralizing / chemistry
-
Antibodies, Neutralizing / immunology
-
Antibodies, Viral / chemistry*
-
Antibodies, Viral / immunology
-
COVID-19 / blood
-
COVID-19 / diagnosis*
-
COVID-19 / epidemiology
-
COVID-19 Serological Testing / methods*
-
COVID-19 Serological Testing / standards
-
Coronavirus Nucleocapsid Proteins / chemistry
-
Coronavirus Nucleocapsid Proteins / immunology*
-
Epidemiological Monitoring
-
Female
-
Humans
-
Immunoglobulin A / chemistry
-
Immunoglobulin A / immunology
-
Immunoglobulin G / chemistry
-
Immunoglobulin G / immunology
-
Immunoglobulin M / chemistry
-
Immunoglobulin M / immunology
-
Male
-
Middle Aged
-
New York / epidemiology
-
Phosphoproteins / chemistry
-
Phosphoproteins / immunology
-
Protein Interaction Domains and Motifs
-
Recombinant Proteins / chemistry
-
Recombinant Proteins / immunology
-
SARS-CoV-2 / classification
-
SARS-CoV-2 / immunology*
-
Sensitivity and Specificity
-
Spike Glycoprotein, Coronavirus / chemistry
-
Spike Glycoprotein, Coronavirus / immunology*
Substances
-
Antibodies, Neutralizing
-
Antibodies, Viral
-
Coronavirus Nucleocapsid Proteins
-
Immunoglobulin A
-
Immunoglobulin G
-
Immunoglobulin M
-
Phosphoproteins
-
Recombinant Proteins
-
Spike Glycoprotein, Coronavirus
-
nucleocapsid phosphoprotein, SARS-CoV-2
-
spike protein, SARS-CoV-2
Grants and funding
Funding for this project was provided by the Cornell University SARS-CoV2 Rapid Research Response Seed Grant CR20, awarded to BW and DGD. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.