CONSTRUCTION OF RECOMBINANT MONOCLONAL ANTIBODY AGAINST HEPATITIS B SURFACE ANTIGEN BY PHAGE DISPLAY

摘要

Single chain antibody fragment (scFv) to surface antigen of Hepatitis B virus was generated by phage display technique. The recombinant protein contains variable regions of murine monoclonal antibody HB3 that recognizes a conformational epitope of hepatis B surface antigen. In this study, immunoglobulin gene was introduced into phagemid vector and rescue by Ml3 helper phage in E. coli TGI. Phage expression and binding activity of recombinant antibody against HBsAg was selected. Infection of E. coli HB2151 by selected recombinant phage resulted in stable, efficient expression of scFv that could be secreted into culture medium. The scFv fragment was purified by affinity chromatography and confirmed by immunoblot analysis, which yield one band of 30 kDa. The activity of recombinant scFv was detected by ELISA, which specifically bound to its cognate antigen at minimum cutoff sensitivity level of 8 ng/ml. These results indicate that specific binding of a scFv to HBsAg can be used for diagnostic purposes. Further experiments, isolated immunoglobulin gene will be cloned into adenovirus vector for transduction of the encoding cDNA into HBV infected hepatocytes and tested its ability to interfere with the HBV replication cycle that might be a novel gene therapy strategy for intracellular immunization against chronic HBV infection.