A SINGLE-CHAIN ANTIBODY FRAGMENT EXPRESSED IN BACTERIA NEUTRALIZES TICK-BORNE FLAVIVIRUSES

A recombinant single chain antibody fragment (scFv) that identifies a neutralizing epitope on the envelope glycoprotein of louping ill (LI) and tick-borne encephalitis (TBE) virus has been developed using a bac teriophage expression system. The mRNA was extracted from a cloned hyb ridoma cell culture that produces a mouse monoclonal antibody (MAb 4.2 ) known to map to amino acids 308-311 of LI and TBE virus, correspondi ng to domain a 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 pill coat protein of filamentous phage. Solid p hase selection of these phage against the LI virus antigen, was necess ary to isolate the correct MAb 4.2 scFv fragment which was subsequentl y 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 infecti vity of louping ill and some other tick-borne flaviviruses. The potent ial of this technique for studying antigen-antibody interactions and f or the development of prophylactic reagents are discussed. (C) 1994 Ac ademic Press, Inc.