title: A single chain antibody fragment expressed in bacteria neutralizes tick-borne flaviviruses.
abstract: A recombinant single chain antibody fragment (scFv) that identifies a neutralizing epitope on the envelope glycoprotein of louping iII (LI) and tick-borne encephalitis (TBE) virus has been developed using a bacteriophage expression system. The mRNA was extracted from a cloned hybridoma cell culture that produces a mouse monoclonal antibody (MAb 4.2) known to map to amino acids 308-311 of LI and TBE virus, corresponding to domain B on the proposed two-dimensional model of the tick-borne encephalitis virus envelope protein. The V-genes encoding the antigen-binding site of MAb 4.2 were amplified and cloned for expression as a fusion protein to the pIII coat protein of filamentous phage. Solid phase selection of these phage against the LI virus antigen, was necessary to isolate the correct MAb 4.2 scFv fragment which was subsequently produced in soluble form in bacteria and harvested from the culture supernatant medium. The characteristics of this expressed single chain antibody were compared with MAb 4.2. The expressed antibody portrayed the antigenic specificity of MAb 4.2 and also neutralized the infectivity of louping iII and some other tick-borne flaviviruses. The potential of this technique for studying antigen-antibody interactions and for the development of prophylactic reagents are discussed.
authors.affiliation: NERC Institute of Virology and Environmental Microbiology, Oxford, United Kingdom.
authors.forename: W
authors.initials: W
authors.lastname: Jiang
authors.forename: T P
authors.initials: TP
authors.lastname: Bonnert
authors.forename: K
authors.initials: K
authors.lastname: Venugopal
authors.forename: E A
authors.initials: EA
authors.lastname: Gould
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics / Q000276:immunology*; D000914:Antibodies, Viral / Q000235:genetics / Q000276:immunology*; D000918:Antibody Specificity; D001483:Base Sequence; D003001:Cloning, Molecular; D004669:Encephalitis Viruses, Tick-Borne / Q000276:immunology*; D004675:Encephalitis, Tick-Borne / Q000517:prevention & control; D004926:Escherichia coli / Q000235:genetics; D005818:Genetic Engineering; D007116:Immunization, Passive; D007135:Immunoglobulin Variable Region / Q000235:genetics / Q000276:immunology*; D051379:Mice; D008969:Molecular Sequence Data; D009500:Neutralization Tests; D011993:Recombinant Fusion Proteins / Q000276:immunology
chemical_list: D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D007135:Immunoglobulin Variable Region; D011993:Recombinant Fusion Proteins