Abstract
SARS-CoV-2 infections continue to spread quickly by human-to-human transmission around the world. Therefore, developing methods to rapidly detect SARS-CoV-2 with high sensitivity are still urgently needed. We produced a monoclonal antibody that specifically detects the N protein of SARS-CoV-2 and recognizes N protein in cell lysates of SARS-CoV-2–infected Vero cells but not in cell lysates of MERS-CoV- or HCoV-OC43-infected Vero cells. This antibody recognized N protein in SARS-CoV-2 clades S, GR, and GH and recognized N protein in all the SARS-CoV-2 clades to ∼300 pfu. Previously, we reported that the coronavirus N protein interacts with the C-terminal domain of the spike protein (Spike CD). In this study, we developed an ELISA-based “bait and prey” system to confirm the interaction between SARS-CoV-2 Spike CD and N protein using recombinant fusion proteins. Furthermore, this system can be modified to quantitatively detect SARS-CoV-2 in culture media of infected cells by monitoring the interaction between the recombinant Spike CD fusion protein and the viral N protein, which is captured by the N protein–specific antibody. Therefore, we conclude that our N protein–specific monoclonal antibody and our ELISA-based bait and prey system could be used to diagnose SARS-CoV-2 infections.
Highlights
Coronaviruses are in the family Coronaviridae and contain genomes composed of positive-sense single-stranded RNA
We produced recombinant MERSCoV N-Bio-His6 protein to analyze the specificity of the SARSCoV-2 N protein-specific monoclonal antibody (Figure 1B)
We formed a complex containing the purified recombinant SARS-CoV-2 N protein and CpG-DNA co-encapsulated in a liposome (DOPE:cholesterol hemisuccinate (CHEMS)) and immunized BALB/c mice with this complex
Summary
Coronaviruses are in the family Coronaviridae and contain genomes composed of positive-sense single-stranded RNA. Severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome coronavirus (MERS-CoV), and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have caused lethal endemics and pandemics in humans. Like SARS-CoV and MERS-CoV, SARS-CoV-2 belongs to the betacoronavirus genus and has a ∼30-kb genome containing genes that encode for structural spike (S), nucleocapsid (N), envelope, and membrane proteins (Khailany et al, 2020). Since the outbreak of coronavirus disease 2019 (COVID-19), caused by SARS-CoV-2 infection, was first reported in December 2019 (Wu and McGoogan, 2020; Zhou P. et al, 2020), the COVID-19 pandemic continues throughout the world, despite the recent start of vaccine administration (WHO, 2021). QRT-PCR is sensitive and is the most specific method for diagnosing COVID-19, this method does not provide rapid results and requires specialized facilities, equipment, and welltrained personnel
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