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Monographic Report
Preparation of human malignant melanoma ganglioside ScFv antibody conjugated quantum dot nanoprobe and its specific binding with human malignant melanoma cells
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Abstract:
Objective:To prepare a nanoprobe, antihuman 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 SDSPAGE. The GD/ScFvMELQDs nanoprobe was prepared by conjugating GD/ScFvMEL antibody with CdTe quantum dot, and its specificity was observed by incubating with MGC803 cells and melanoma A375 cells. Results:The recombinant pET32aGD/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 SDSPAGE. The purified and refoldedGD/ScFvMEL antibody was effectively conjugated with CdTe quantum dot, and the resulting GD/ScFvMELQDs nanoprobe was successfully prepared. The GD/ScFvMELQDs nanoprobe could specifically bind melanoma A375 cells, but could not bind stomach cancer MGC803 cells. Conclusion:We have successfully prepared an antihuman melanoma ganglioside singlechain antibodyCdTe quantum dot nanoprobe, which can specifically bind melanoma cells.
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Project Supported:
Project supported by the National High Technology Research and Development Program (863) of China (No. 2007AA022004), and the Major State Basic Research Development Program (973) of China (No. 2010CB933901)
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