目的：制备抗人黑素瘤细胞单链抗体与碲化镉量子点(CdTe guantun dot, CdTe)连接的纳米探针，观察其对恶性黑素瘤细胞的特异性结合效果。方法：将抗人黑素瘤神经节苷脂单链抗体（antihuman melanoma ganglioside single chain variable fragment antibody，GD/ScFvMEL）基因克隆到pET32a（+）载体中，然后在BL21（DE3）细菌中进行诱导表达，采用变性法纯化表达的蛋白，再用改良的透析法复性目的蛋白，SDSPAGE分析表达的GD/ScFvMEL抗体蛋白。制备的GD/ScFvMEL抗体与量子点连接制备成纳米探针GD/ScFvMELQDs，然后与黑素瘤A375细胞株及胃癌MGC803细胞株孵育，观察纳米探针与肿瘤细胞结合的特异性。结果：PCR、双酶切和序列测定证实重组子的拼接完全正确，SDSPADE结果显示，GD/ScFvMEL抗体的表达量达40%。纯化复性后的GD/ScFvMEL抗体连接量子点制备GD/ScFvMELQDs纳米探针，经扫描电镜、X衍射分析、荧光光谱和SDSPAGE分析证实纳米探针的成功制备。GD/ScFvMELQDs纳米探针能特异性结合A375黑素瘤细胞，不与胃癌细胞MGC803细胞结合。结论：成功制备抗人黑素瘤神经节苷脂单链抗体量子点纳米探针，该探针可特异性结合黑素瘤细胞。

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.

国家高技术研究发展（863）计划重点项目（No. 2007AA022004）；国家重点基础研究发展（973）计划资助项目（No. 2010CB933900）