Preparation of highly specific monoclonal antibodies against SARS‐CoV‐2 nucleocapsid protein and the preliminary development of antigen detection test strips

• Published in: Journal of medical virology 2022-04, Vol.94 (4), p.1633-1640
• Main Authors: Xie, Chengzuo, Ding, Haojie, Ding, Jianzu, Xue, Yangji, Lu, Shaohong, Lv, Hangjun
• Format: Article
• Language: English
• Subjects: Animals; Antibodies, Monoclonal - immunology; Antigens; Ascites; colloidal gold immunochromatography; Coronavirus Nucleocapsid Proteins - blood; Coronavirus Nucleocapsid Proteins - genetics; Coronavirus Nucleocapsid Proteins - immunology; Coronaviruses; COVID-19; COVID-19 - blood; COVID-19 - diagnosis; COVID-19 Serological Testing - instrumentation; COVID-19 Serological Testing - methods; Delta (B.1.617.2) variant; E coli; Humans; Immunoassay; Immunoregulation; Life cycles; Mice; Monoclonal antibodies; Mutation; N gene; N protein; Nucleocapsids; Phosphoproteins - blood; Phosphoproteins - genetics; Phosphoproteins - immunology; Proteins; Recombinant Proteins - genetics; Recombinant Proteins - immunology; recombinant SARS‐CoV‐2 nucleocapsid protein; Respiratory diseases; SARS-CoV-2 - genetics; SARS-CoV-2 - immunology; SARS-CoV-2 - isolation & purification; SARS‐CoV‐2; Sensitivity and Specificity; Severe acute respiratory syndrome; Severe acute respiratory syndrome coronavirus 2; the monoclonal antibodies; Therapeutic targets; Viral diseases; Virology
• ISSN: 0146-6615; 1096-9071
• DOI: 10.1002/jmv.27520

The coronavirus disease 2019 (COVID‐19) is outbreaking all over the world. To help fight this disease, it is necessary to establish an effective and rapid detection method. The nucleocapsid (N) protein of severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) is involved in viral replication, assembly, and immune regulation and plays an important role in the viral life cycle. Moreover, the N protein also could be a diagnostic factor and potential drug target. Therefore, by synthesizing the N gene sequence of SARS‐CoV‐2, constructing the pET‐28a (+)‐N recombinant plasmid, we expressed the N protein in Escherichia coli and obtained 15 monoclonal antibody (mAbs) against SARS‐CoV‐2‐N protein by the hybridomas and ascites, then an immunochromatographic test strip method detecting N antigen was established. In this study, we obtained 14 high‐titer and high‐specificity monoclonal antibodies, and the test strips exclusively react with the SARS‐CoV‐2‐N protein and no cross‐reactivity with other coronavirus and also recognize the recombinant N protein of Delta (B.1.617.2) variant. These mAbs can be used for the early and rapid diagnosis of SARS‐CoV‐2 infection through serological antigen.