Generation of monoclonal antibodies against SARS-CoV-2

Abstract

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is the causative agent of the Coronavirus disease 2019 (COVID-19). As of 14th December 2021, COVID-19 pandemic has caused over two hundred and seventy million of laboratory-confirmed cases and over five million of deaths exerting a heavy burden on public health worldwide. Generation of specific antibodies recognizing antigens of SARS-CoV-2 is urgently needed for studying and detecting SARS-CoV-2 infection. In this work, I produced a recombinant nucleocapsid protein (NP) of SARS-CoV-2 by using a bacterial overexpression system. Three mice were immunized by this purified recombinant NP and their antibody levels were monitored. Upon the completion of the immunization scheme, the antibody producing cells from the immunized mice were isolated and then immortalized by fusing with a myeloma cell line, NS1, using the standard hybridoma technique. After primary screening and subcloning, I characterized the specificity and the sensitivity of each monoclonal antibody by using Western blot analysis, immunofluorescent staining, and enzyme-linked immunosorbent assay (ELISA). In total, I generated seven hybridomas which produced antibodies with high specificity and sensitivity. By using one of my purified antibodies, we were able to discover a new cell entry mechanism of SARS-CoV-2 by monitoring the early step of SARS-CoV-2 infection. I also explored the feasibility of utilizing these antibodies in the development of antibody-based diagnostic approaches and treatment options for COVID-19.