State Key Laboratory of Electroanalytical Chemistry, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun 130022, PR China.
State Key Laboratory of Electroanalytical Chemistry, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun 130022, PR China; University of Chinese Academy of Sciences, No.19A Yuquan Road, Beijing 100049, PR China.
Talanta. 2018 Feb 1;178:116-121. doi: 10.1016/j.talanta.2017.09.034. Epub 2017 Sep 13.
Telomerase is extensively expressed in various cancer cells and recognized as a target for cancer drug discovery. In the present study, a simple and amplification-free fluorescence assay based on polyamidoamine starburst dendrimer (PAMAM dendrimer)-activated paper device is proposed for sensitive detection of telomerase activity through hybridization of Cy5 modified single strand DNA probes with telomerase extension products. The paper substrate is fabricated by hand drawing according to a template, which is low cost, instrument free and easy operation. PAMAM is rich in amino groups on its surface and employed to immobilize the telomerase substrate (TS) primer. Highly sensitive detection of telomerase activity in HeLa cell lysate of 10 cells is achieved since the PAMAM dendrimer-activated paper surface can provide high density of binding sites for immobilization of TS primer. The experimental results also demonstrate that the assay can be employed to evaluate telomerase activity levels of various cell lines and screen telomerase inhibitors.
端粒酶在各种癌细胞中广泛表达,被认为是癌症药物发现的靶点。在本研究中,提出了一种基于聚酰胺-胺星状树枝状大分子(PAMAM 树枝状大分子)激活纸基器件的简单、无需扩增的荧光分析方法,通过 Cy5 修饰的单链 DNA 探针与端粒酶延伸产物的杂交,实现了端粒酶活性的灵敏检测。纸基是根据模板手工绘制的,成本低、无仪器且易于操作。PAMAM 表面富含氨基,用于固定端粒酶底物(TS)引物。由于 PAMAM 树枝状大分子激活的纸表面可以为 TS 引物的固定提供高密度的结合位点,因此可以实现对 10 个细胞的 HeLa 细胞裂解物中端粒酶活性的高灵敏度检测。实验结果还表明,该测定法可用于评估各种细胞系中端粒酶活性水平,并筛选端粒酶抑制剂。