School of Pharmaceutical Sciences, Shandong University, 250012 Jinan, PR China.
Key Laboratory for Colloid and Interface Chemistry of Education Ministry, School of Chemistry and Chemical Engineering, Shandong University, 250100 Jinan, PR China.
Talanta. 2018 Feb 1;178:458-463. doi: 10.1016/j.talanta.2017.09.057. Epub 2017 Sep 19.
Human telomerase is an endogenous ribonucleoprotein that is over-expressed in most types of malignant cancer cells. Sensitive and specific detection of telomerase activity is crucial for better understanding its role in cancer cells and further exploring its function in cancer diagnosis. Here, we develop convertible DNA ends-based silver nanoprobes for sensitive and specific colorimetric detection telomerase activity. Silver nanoprobes are constructed by modifying telomerase binding substrates (TS) that are pre-hybridized with complementary sequences onto silver nanoparticles (AgNPs), via the coordination between consecutive cytosines in TS strand and AgNPs. This forms blunt-end terminated, double-stranded DNA on the surface of AgNPs. Under the action of telomerase, TS on the silver nanoprobes are elongated with telomeric repeats, converting DNA stiff blunt ends to flexible single-stranded dangling ends. The dangling ends enhance the stability of nanoprobes and relieve their salt-induced aggregation, and the solution shows a yellow color. When telomerase is inactive, the blunt end-terminated nanoprobes cannot resist salt-induced aggregation, resulting in a gray color of solution. Based on telomerase-regulated DNA "blunt-dangling" ends conversion-induced AgNPs' dispersity and color change, colorimetric detection of the endogenous telomerase with AgNPs is realized. The detection limit is equivalent to 1 cell/μL of telomerase activity, and extracts from cancer cells and normal cells are visually distinguished through color difference. The proposed strategy will offer a new approach for reliable, convenient quantification of telomerase activity in biochemical research and clinical diagnosis.
人端粒酶是一种内源性的核蛋白，在大多数类型的恶性癌细胞中过度表达。端粒酶活性的敏感和特异性检测对于更好地理解其在癌细胞中的作用以及进一步探索其在癌症诊断中的功能至关重要。在这里，我们开发了基于可转换 DNA 末端的银纳米探针，用于敏感和特异性比色法检测端粒酶活性。银纳米探针通过将与互补序列预先杂交的端粒酶结合底物（TS）修饰在银纳米粒子（AgNPs）上而构建，通过 TS 链中连续胞嘧啶与 AgNPs 之间的配位。这在 AgNPs 表面形成末端平齐的双链 DNA。在端粒酶的作用下，银纳米探针上的 TS 被端粒重复序列延伸，将 DNA 硬平齐末端转换为灵活的单链悬空末端。悬空末端增强了纳米探针的稳定性并减轻了盐诱导的聚集，溶液呈黄色。当端粒酶失活时，末端平齐的纳米探针不能抵抗盐诱导的聚集，导致溶液呈灰色。基于端粒酶调节的 DNA“平齐-悬空”末端转换诱导的 AgNPs 分散性和颜色变化，实现了基于 AgNPs 的内源性端粒酶的比色检测。检测限相当于 1 个细胞/μL 的端粒酶活性，并且可以通过色差目视区分癌细胞和正常细胞的提取物。所提出的策略将为生化研究和临床诊断中可靠、方便地定量端粒酶活性提供一种新方法。
