[Construction and expression of mouse-human chimeric Fab fragment gene of monoclonal antibody SZ-2 against platelet]

Abstract

Aim: To reduce immunogenicity of a monoclonal antibody SZ-2 specific for human platelet.

Methods: Reverse transcription and polymerize chain reaction were used to amplify the variable region genes of monoclonal antibody SZ-2. The cloned V(H) and V(L) genes were sequenced and fused to human IgG1 constant region gene CH1 and Ckappa in plasmid pSW1. The recombinant plasmid were transformed into E. coli. The expressed recombinant proteins were analysed.

Results: The V(H) and V(L) genes were homologous with the published gene sequences of mouse antibody variable region. The concentration of chimeric Fab fragment in expression supernatant was about 180 microg/L detected by ELISA. Western blot analysis showed that SZ-2 Fab/Hu maintained the binding activity to human platelet GPIb. The recombinant proteins could suppress platelet aggregation induced by Ristocatin.

Conclusion: The variable region genes of SZ-2 are cloned and the mouse-human chimeric Fab fragment is expressed successfully in E. coli.