Abstract: To construct a bispecific single chain antibody fragment antiCD3×antiTP3 diabody with dual specificity for both the human osteosarcoma cell antigen TP3 and CD3εchain of CD3/TCR. Methods： The expression plasmid pKIDCD3 × TP3 was constructed with gene of antibody variable fragment, and then it was transinfected into the E. coli K12 strain XL1Blue to obtain diabody. The diabody was purified by IMAC and its affinity was determined by flow cytometry and PAGE, at last, it was incubated with human T cell and osteosarcoma cell. Results： The diabody was absolute and stable in human plasma with high specificity for human T cells and osteosarcoma cells, and it also makes T cells approach target cells and lyse them. Conclusion： The diabody not only stimulates and retargets CTLs to osteosarcoma cells but also lyses them.

Key words: Osteosarcoma, T lymphocyte, Monoclonal antibodies, Singlechain Fv, Diabody, Cellular immunity