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Design and Development of an Antigen Test for SARS-CoV-2 Nucleocapsid Protein to Validate the Viral Quality Assurance Panels.
Ray, Partha; Ledgerwood-Lee, Melissa; Brickner, Howard; Clark, Alex E; Garretson, Aaron; Graham, Rishi; Van Zant, Westley; Carlin, Aaron F; Aronoff-Spencer, Eliah S.
Afiliação
  • Ray P; Department of Medicine, Division of Infectious Diseases and Global Public Health, University of California, San Diego, CA 92093, USA.
  • Ledgerwood-Lee M; Department of Medicine, Division of Infectious Diseases and Global Public Health, University of California, San Diego, CA 92093, USA.
  • Brickner H; Department of Medicine, Division of Infectious Diseases and Global Public Health, University of California, San Diego, CA 92093, USA.
  • Clark AE; Department of Medicine, Division of Infectious Diseases and Global Public Health, University of California, San Diego, CA 92093, USA.
  • Garretson A; Department of Medicine, Division of Infectious Diseases and Global Public Health, University of California, San Diego, CA 92093, USA.
  • Graham R; Department of Medicine, Division of Infectious Diseases and Global Public Health, University of California, San Diego, CA 92093, USA.
  • Van Zant W; Department of Medicine, Division of Infectious Diseases and Global Public Health, University of California, San Diego, CA 92093, USA.
  • Carlin AF; Department of Medicine, Division of Infectious Diseases and Global Public Health, University of California, San Diego, CA 92093, USA.
  • Aronoff-Spencer ES; Department of Pathology, University of California, San Diego, CA 92093, USA.
Viruses ; 16(5)2024 04 24.
Article em En | MEDLINE | ID: mdl-38793544
ABSTRACT
The continuing mutability of the SARS-CoV-2 virus can result in failures of diagnostic assays. To address this, we describe a generalizable bioinformatics-to-biology pipeline developed for the calibration and quality assurance of inactivated SARS-CoV-2 variant panels provided to Radical Acceleration of Diagnostics programs (RADx)-radical program awardees. A heuristic genetic analysis based on variant-defining mutations demonstrated the lowest genetic variance in the Nucleocapsid protein (Np)-C-terminal domain (CTD) across all SARS-CoV-2 variants. We then employed the Shannon entropy method on (Np) sequences collected from the major variants, verifying the CTD with lower entropy (less prone to mutations) than other Np regions. Polyclonal and monoclonal antibodies were raised against this target CTD antigen and used to develop an Enzyme-linked immunoassay (ELISA) test for SARS-CoV-2. Blinded Viral Quality Assurance (VQA) panels comprised of UV-inactivated SARS-CoV-2 variants (XBB.1.5, BF.7, BA.1, B.1.617.2, and WA1) and distractor respiratory viruses (CoV 229E, CoV OC43, RSV A2, RSV B, IAV H1N1, and IBV) were assembled by the RADx-rad Diagnostics core and tested using the ELISA described here. The assay tested positive for all variants with high sensitivity (limit of detection 1.72-8.78 ng/mL) and negative for the distractor virus panel. Epitope mapping for the monoclonal antibodies identified a 20 amino acid antigenic peptide on the Np-CTD that an in-silico program also predicted for the highest antigenicity. This work provides a template for a bioinformatics pipeline to select genetic regions with a low propensity for mutation (low Shannon entropy) to develop robust 'pan-variant' antigen-based assays for viruses prone to high mutational rates.
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Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Fosfoproteínas / Proteínas do Nucleocapsídeo de Coronavírus / SARS-CoV-2 / COVID-19 / Antígenos Virais Limite: Animals / Humans Idioma: En Revista: Viruses Ano de publicação: 2024 Tipo de documento: Article País de afiliação: Estados Unidos

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Fosfoproteínas / Proteínas do Nucleocapsídeo de Coronavírus / SARS-CoV-2 / COVID-19 / Antígenos Virais Limite: Animals / Humans Idioma: En Revista: Viruses Ano de publicação: 2024 Tipo de documento: Article País de afiliação: Estados Unidos