Antigenic properties of the SARS-CoV-2 nucleoprotein are altered by the RNA admixture

• Published in: PeerJ (San Francisco, CA) Vol. 10; p. e12751
• Main Authors: Kolesov, Denis E, Sinegubova, Maria V, Safenkova, Irina V, Vorobiev, Ivan I, Orlova, Nadezhda A
• Format: Journal Article
• Language: English
• Published: United States PeerJ. Ltd 07-01-2022; PeerJ, Inc; PeerJ Inc
• Subjects: Analysis; Antibodies; Antibodies, Viral; Antigens; Biotechnology; Chromatography; Coronaviruses; COVID-19; COVID-19 vaccines; Cytoplasm; Disease transmission; E coli; ELISA; Enzyme-linked immunosorbent assay; Escherichia coli; Escherichia coli - genetics; Flow velocity; Health aspects; Humans; Infections; IVD; Molecular Biology; Nucleoprotein; Nucleoproteins; Pandemics; Plasmids; Proteins; Purification; Ribonucleic acid; RNA; SARS-CoV-2; SARS-CoV-2 - genetics; Scientific equipment and supplies industry; Serological testing; Serology; Severe acute respiratory syndrome coronavirus 2; Sodium chloride; Viral antibodies; Virology; United States; Germany; Russia; United States > US; Nucleoprotein; SARS-CoV-2; Serological testing; IVD; ELISA
• ISSN: 2167-8359; 2167-8359
• DOI: 10.7717/peerj.12751

Determining the presence of antibodies to the SARS-CoV-2 antigens is the best way to identify infected people, regardless of the development of symptoms of COVID-19. The nucleoprotein (NP) of the SARS-CoV-2 is an immunodominant antigen of the virus; anti-NP antibodies are detected in persons previously infected with the virus with the highest titers. Many test systems for detecting antibodies to SARS-CoV-2 contain NP or its fragments as antigen. The sensitivity and specificity of such test systems differ significantly, which can be explained by variations in the antigenic properties of NP caused by differences in the methods of its cultivation, isolation and purification. We investigated this effect for the
-derived SARS-CoV-2 NP, obtained from the cytoplasm in the soluble form. We hypothesized that co-purified nucleic acids that form a strong complex with NP might negatively affect NP's antigenic properties. Therefore, we have established the NP purification method, which completely eliminates the RNA in the NP preparation. Two stages of RNA removal were used: treatment of the crude lysate of 
with RNase A and subsequent selective RNA elution with 2 M NaCl solution. The resulting NP without RNA has a significantly better signal-to-noise ratio when used as an ELISA antigen and tested with a control panel of serum samples with antibodies to SARS-CoV-2; therefore, it is preferable for in vitro diagnostic use. The same increase of the signal-to-noise ratio was detected for the free N-terminal domain of the NP. Complete removal of RNA complexed with NP during purification will significantly improve its antigenic properties, and the absence of RNA in NP preparations should be controlled during the production of this antigen.