Expression and Characterization of Recombinant, Tetrameric and Enzymatically Active Influenza Neuraminidase for the Setup of an Enzyme-Linked Lectin-Based Assay

• Published in: PloS one 2015-08, Vol.10 (8), p.e0135474-e0135474
• Main Authors: Prevato, Marua, Ferlenghi, Ilaria, Bonci, Alessandra, Uematsu, Yasushi, Anselmi, Giulia, Giusti, Fabiola, Bertholet, Sylvie, Legay, Francois, Telford, John Laird, Settembre, Ethan C, Maione, Domenico, Cozzi, Roberta
• Format: Article
• Language: English
• Subjects: Animal experimentation; Animals; Antibodies, Viral - immunology; Antibody Formation - immunology; Antigens; Antigens, Viral - immunology; Antiviral agents; Avian influenza; Birds; Cell Line; Chemical properties; Cross Protection - immunology; Cross Reactions - immunology; Cross-protection; Design; Development and progression; Diagnostic software; Diagnostic systems; Disease control; Enzyme-Linked Immunospot Assay - methods; Enzymes; Exo-a-sialidase; Female; Gene expression; Hemagglutinin Glycoproteins, Influenza Virus - immunology; Hemagglutinins; Immunity; Immunogenicity; Immunoglobulins; Immunology; Infections; Influenza; Influenza A Virus, H1N1 Subtype - immunology; Influenza A Virus, H5N1 Subtype - immunology; Influenza in Birds - immunology; Influenza vaccines; Influenza Vaccines - immunology; Lectins; Lectins - immunology; Livestock; Mammalian cells; Mice; Microscopy; Molecular biology; Neuraminidase - immunology; Orthomyxoviridae Infections - immunology; Pandemics; R&D; Recombinant Proteins - immunology; Research & development; Ribonucleic acid; RNA; Swine; Swine flu; Vaccines; Viral infections; Viral Proteins - immunology; Virology; Viruses
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0135474

Developing a universal influenza vaccine that induces broad spectrum and longer-term immunity has become an important potentially achievable target in influenza vaccine research and development. Hemagglutinin (HA) and neuraminidase (NA) are the two major influenza virus antigens. Although antibody responses against influenza virus are mainly directed toward HA, NA is reported to be more genetically stable; hence NA-based vaccines have the potential to be effective for longer time periods. NA-specific immunity has been shown to limit the spread of influenza virus, thus reducing disease symptoms and providing cross-protection against heterosubtypic viruses in mouse challenge experiments. The production of large quantities of highly pure and stable NA could be beneficial for the development of new antivirals, subunit-based vaccines, and novel diagnostic tools. In this study, recombinant NA (rNA) was produced in mammalian cells at high levels from both swine A/California/07/2009 (H1N1) and avian A/turkey/Turkey/01/2005 (H5N1) influenza viruses. Biochemical, structural, and immunological characterizations revealed that the soluble rNAs produced are tetrameric, enzymatically active and immunogenic, and finally they represent good alternatives to conventionally used sources of NA in the Enzyme-Linked Lectin Assay (ELLA).