A Reliable Indirect ELISA Protocol for Detection of Human Antibodies Directed to SARS-CoV-2 NP Protein

• Main Authors: Arwa A. Faizo, Thamir A. Alandijany, Ayman T. Abbas, Sayed S. Sohrab, Sherif A. El-Kafrawy, Ahmed M. Tolah, Ahmed M. Hassan, Esam I. Azhar
• Format: Article
• Language: English
• Published: MDPI AG 2021-05-01
• Series: Diagnostics
• Subjects: COVID-19; SARS-CoV-2; ELISA; serology; seroprevalence; nucleocapsid
• Online Access: https://www.mdpi.com/2075-4418/11/5/825

A few months ago, the availability of a reliable and cost-effective testing capacity for COVID-19 was a concern for many countries. With the emergence and circulation of new SARS-CoV-2 variants, another layer of challenge can be added for COVID-19 testing at both molecular and serological levels. This is particularly important for the available tests principally designed to target the S gene/protein where multiple mutations have been reported. Herein, the SARS-CoV-2 NP recombinant protein was utilized to develop a simple and reliable COVID-19 NP human IgG ELISA. The optimized protocol was validated against a micro-neutralization (MN) assay, in-house S-based ELISA, and commercial chemiluminescence immunoassay (CLIA). The developed assay provides 100% sensitivity, 98.9% specificity, 98.9% agreement, and high overall accuracy with an area under curve equal to 0.9998 ± 0.0002 with a 95% confidence interval of 0.99 to 1.00. The optical density values of positive samples significantly correlated with their corresponding MN titers. The assay specifically detects IgG antibodies to the SARS-CoV-2 NP protein and does not cross-detect IgG to the viral S protein. Moreover, it does not cross-react with antibodies related to other coronaviruses (e.g., the Middle East respiratory syndrome coronavirus or human coronavirus HKU1). The availability of this reliable COVID-19 NP IgG ELISA protocol is highly valuable for its diagnostic and epidemiological applications.