title: Isolation and characterisation of Ebolavirus-specific recombinant antibody fragments from murine and shark immune libraries.
abstract: Members of the genus Ebolavirus cause fulminating outbreaks of disease in human and non-human primate populations with a mortality rate up to 90%. To facilitate rapid detection of these pathogens in clinical and environmental samples, robust reagents capable of providing sensitive and specific detection are required. In this work recombinant antibody libraries were generated from murine (single chain variable domain fragment; scFv) and nurse shark, Ginglymostoma cirratum (IgNAR V) hosts immunised with Zaire ebolavirus. This provides the first recorded IgNAR V response against a particulate antigen in the nurse shark. Both murine scFv and shark IgNAR V libraries were panned by phage display technology to identify useful antibodies for the generation of immunological detection reagents. Two murine scFv were shown to have specificity to the Zaire ebolavirus viral matrix protein VP40. Two isolated IgNAR V were shown to bind to the viral nucleoprotein (NP) and to capture viable Zaire ebolavirus with a high degree of sensitivity. Assays developed with IgNAR V cross-reacted to Reston ebolavirus, Sudan ebolavirus and Bundibugyo ebolavirus. Despite this broad reactivity, neither of IgNAR V showed reactivity to Côte d'Ivoire ebolavirus. IgNAR V was substantially more resistant to irreversible thermal denaturation than murine scFv and monoclonal IgG in a comparative test. The demonstrable robustness of the IgNAR V domains may offer enhanced utility as immunological detection reagents in fieldable biosensor applications for use in tropical or subtropical countries where outbreaks of Ebolavirus haemorrhagic fever occur.
authors.affiliation: Defence Science and Technology Laboratory, Porton Down, Salisbury SP4 0JQ, UK. sagoodchild@mail.dstl.gov.uk
authors.forename: Sarah A
authors.initials: SA
authors.lastname: Goodchild
authors.forename: Helen
authors.initials: H
authors.lastname: Dooley
authors.forename: Randal J
authors.initials: RJ
authors.lastname: Schoepp
authors.forename: Martin
authors.initials: M
authors.lastname: Flajnik
authors.forename: Stephen G
authors.initials: SG
authors.lastname: Lonsdale
mesh_terms: D000818:Animals; D000918:Antibody Specificity / Q000276:immunology*; D015153:Blotting, Western; D029043:Ebolavirus / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D007128:Immunoglobulin Fragments / Q000276:immunology / Q000302:isolation & purification*; D051379:Mice; D019151:Peptide Library; D011489:Protein Denaturation; D011994:Recombinant Proteins / Q000276:immunology / Q000302:isolation & purification; D012754:Sharks / Q000276:immunology*; D014764:Viral Proteins / Q000276:immunology*
chemical_list: D007128:Immunoglobulin Fragments; D019151:Peptide Library; D011994:Recombinant Proteins; D014764:Viral Proteins