Qiu Suyan, Zhao Fusheng, Zenasni Oussama, Li Jingting, Shih Wei-Chuan
Department of Electrical and Computer Engineering, University of Houston, 4800 Calhoun Road, Houston, TX 77204, USA.
Nanoscale Horiz. 2017 Jul 1;2(4):217-224. doi: 10.1039/c7nh00042a. Epub 2017 May 31.
Telomerase, an enzyme known to catalyze telomere elongation by adding TTAGGG [thymine (T), adenine (A), and guanine (G)] repeats to the end of telomeres, is vital for cell proliferation. Overexpression of telomerase has been found in most tumor cells, resulting in telomere dysfunction and uncontrolled cellular proliferation. Thus, telomerase has been considered as a potential cancer biomarker, as well as a potential target in cancer therapy. In this study, telomerase-catalyzed growth of tandem G-quadruplex (G4) assembled on a nanoporous gold array (NPGA) resulted in the formation of three-dimensional hybrid nanoarchitectures. The generated nanostructure then captured malachite green (MG) (reporter molecule) without the need of a complicated labeling process. Upon laser irradiation, the captured MG molecules produced a surface-enhanced Raman scattering (SERS) signal that was generated by an abundant amount of plasmonic hot spots in the NPGA substrates. A limit of detection (LOD) of 10 IU along with a linear range, which was 3 orders of magnitude, was achieved, which was equivalent to the telomerase amount extracted from 20 HeLa cells. The LOD is 2 orders of magnitude better than that of the commercial enzyme-linked immunosorbent assay (ELISA), and it approaches that of the most sensitive technique, telomeric repeat amplification protocols (TRAP), which require a laborious and equipment-intensive polymerase chain reaction (PCR). In addition, X-ray photoelectron spectroscopy (XPS) was used to chemically identify and quantify the telomerase activity on the sensitized NPGA surface. Furthermore, the sensor was applied to screen the effectiveness of anti-telomerase drugs such as zidovudine, thus demonstrating the potential use of the sensor in telomerase-based diagnosis and drug development. Moreover, the framework represents a novel paradigm of collaborative plasmonic intensification and catalytic multiplication (c-PI/CM) for label-free biosensing.
端粒酶是一种通过在端粒末端添加TTAGGG（胸腺嘧啶（T）、腺嘌呤（A）和鸟嘌呤（G））重复序列来催化端粒延长的酶，对细胞增殖至关重要。在大多数肿瘤细胞中都发现了端粒酶的过表达，导致端粒功能障碍和细胞增殖失控。因此，端粒酶被认为是一种潜在的癌症生物标志物，也是癌症治疗的潜在靶点。在本研究中，端粒酶催化在纳米多孔金阵列（NPGA）上组装的串联G-四链体（G4）生长，导致三维混合纳米结构的形成。然后，生成的纳米结构无需复杂的标记过程就能捕获孔雀石绿（MG）（报告分子）。在激光照射下，捕获的MG分子产生了由NPGA底物中大量等离子体热点产生的表面增强拉曼散射（SERS）信号。实现了10 IU的检测限以及3个数量级的线性范围，这相当于从20个HeLa细胞中提取的端粒酶量。该检测限比商业酶联免疫吸附测定（ELISA）的检测限好2个数量级，并且接近最灵敏技术端粒重复序列扩增方案（TRAP）的检测限，而TRAP需要繁琐且设备密集的聚合酶链反应（PCR）。此外，X射线光电子能谱（XPS）用于化学鉴定和定量敏化NPGA表面的端粒酶活性。此外，该传感器被用于筛选抗端粒酶药物如齐多夫定的有效性，从而证明了该传感器在基于端粒酶的诊断和药物开发中的潜在用途。此外，该框架代表了一种用于无标记生物传感的协同等离子体增强和催化倍增（c-PI/CM）的新范例。
