Preparation of human malignant melanoma ganglioside ScFv antibody-conjugated quantum dot nanoprobe and its specific binding with human malignant melanoma cells / 中国肿瘤生物治疗杂志

ABSTRACT

Objective: To prepare a nanoprobe, anti-human melanoma ganglioside single chain variable fragment (GD/ScFvMEL) antibody conjugated with CdTe quantum dot, and to observe its ability to specifically bind human malignant melanoma cells. Methods: The GD/ScFvMEL gene was cloned into pET32a (+), and the plasmid was then transformed into E. coli BL21 (DE3) for GD/ScFvMEL protein antibody expression. The expressed GD/ScFvMEL antibody was purified by denaturing method and further refolded by modified dialysis method. The purified GD/ScFvMEL antibody was analyzed by SDS-PAGE. The GD/ScFvMEL-QDs nanoprobe was prepared by conjugating GD/ScFvMEL antibody with CdTe quantum dot, and its specificity was observed by incubating with MGC-803 cells and melanoma A375 cells. Results: The recombinant pET32a-GD/ScFvMEL was constructed and confirmed by PCR, restriction endonuclease analysis and DNA sequencing. The proportion of expressed GD/ScFvMEL antibody in total bacteria proteins was about 40% as detected by SDS-PAGE. The purified- and refolded-GD/ScFvMEL antibody was effectively conjugated with CdTe quantum dot, and the resulting GD/ScFvMEL-QDs nanoprobe was successfully prepared. The GD/ScFvMEL-QDs nanoprobe could specifically bind melanoma A375 cells, but could not bind stomach cancer MGC-803 cells. Conclusion: We have successfully prepared an anti-human melanoma ganglioside single-chain antibody-CdTe quantum dot nanoprobe, which can specifically bind melanoma cells.