Expression and functional analysis of fusion protein for the human anti-HBsAg scFv antibody fragment-consensus interferon
- VernacularTitle:人源抗HBsAgscFv与重组复合干扰素融合蛋白的高效表达及活性鉴定
- Author:
Shunai LIU
;
Asano RYUTARO
;
Xue WANG
;
Kumagai IZUMI
;
Kudo TOSHIO
;
Daozhen XU
- Publication Type:Journal Article
- Keywords:
HBsAg;
Human scFv;
cIFN;
Fusion protein;
Bacterial expression;
Activity
- From:
Chinese Journal of Immunology
2000;0(09):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective:Express a human anti-HBsAg single chain antibody fragment(scFv)-consensus interferon (cIFN) fusion protein by bacterial expression system and analyse the function of the fusion protein.Methods:Human anti-HBsAg single chain monoclonal antibody cDNA encoding the variable regions of immunoglobulin from PBMC of Hepatitis B patient. Consensus interferon gene was produced by overlap PCR.A plasmid for production of cIFN-scFV fusion protein was constructed, then the expression vector pRA cIFN-scFV transformed with the E.coli strain BL21(DE3). The gene product was analysed SDS-PAGE and Western blotting, then was solubilized by guanidine hydyochloride, refolded by conventional dilution method, and purified using metal-chelating chromatography. The immune and functional analysis of the resulting fusion protein have been studied by ELISA,FACS(Flow cytometry),MTS assay and hemaglutination inhibition test.Results:The authors isolated and characterized the human anti-HBsAg single chain antibody fragment(scFv)-consensus interferon (cIFN) fusion protein. The resulting human anti-HBsAg scFv-cIFN fusion protein was bound to react with HBsAg and cIFN, this react show that highly specific and bioactivity.Conclusion:A human anti-HBsAg single chain antibody fragment(scFv)-consensus interferon (cIFN) fusion protein was produced by bacterial expression system in this study. This genetically engineered human anti-HBsAg scFv-cIFN fusion protein promises to be an important reagent for hepatitis B immunotherapy.
