Construction And Experimental Targeting Studies Of A Single Chain Fv Immunotoxin Against Human Hepatocellular Carionoma

The murine monoclonal antibody (IgG1 k, HAb25) was rasied against human hepatocellular carcinoma. Here four experiments were made in order to identify the specificity of the antibody and to construct a single chain Fv immunotoxin derived from it. Firstly the radioimuno imagings of ¹³¹I-HAb25 for the nude mice bearing xenografts of HCC and the paients with HCC were taken; Conjugating the HAb25 with ADM was performed the cytotoxicity in vitro and targeing therapy in the nude mice bearing xenografts of HCC in vivo. The results revealed that the HAb25 was able to target in vitro and in vivo and may be useful for clinical application. Secondly the heavy and light variable regiongenes were isolated by RT-PCR technique from the HAb25 hybridoma; The sequence analysis of the variable region genes showed that they were new and rearrenged variable region genes. Thirdly the VH and VL genes were connected by a (Gly4Ser)3 polypeptide linker to form HAb25 (ScFv) antibody gene; The constructed HAb25 (ScFv) antibody gene was high expressed as a form of fusion protein, which retained comparatively the specificity and affinity of its native antibody. Finally a ScFv immnotoxin was made by fusing the HAb25 (ScFv) domain to PE40,a...