Key Laboratory of Analytical Chemistry for Biology and Medicine (Ministry of Education), College of Chemistry and Molecular Sciences, Research Center for Nanobiology and Nanomedicine (MOE 985 Innovative Platform), Wuhan Institute of Biotechnology, Wuhan University, Wuhan, 430072, PR China.
Biosens Bioelectron. 2011 Nov 15;29(1):46-52. doi: 10.1016/j.bios.2011.07.064. Epub 2011 Aug 2.
In this work, we have developed a simple and sensitive method for ATP detection using silica nanoparticles (NPs) as the platform and hoechst33258 as the signal reporter. The ATP-binding aptamers hybridize with the probe DNA (DNA(p)) immobilized NPs to form the aptamer/DNA(p) duplex on the NPs surface. The conformational change of the aptamer leads to the decrease of the aptamer/DNA(p) duplex on the NPs due to the ATP-binding aptamer switches its structure from the aptamer/DNA(p) duplex to the aptamer/target complex in the presence of ATP. ATP detection can be easily realized by separating the silica nanoparticles and adding the hoechst33258 of intercalating to aptamer/DNA(p) (dsDNA). Good selectivity between ATP and CTP, GTP or UTP has been demonstrated, which is due to the specific recognition between ATP aptamer and ATP. The K(d) was estimated to be ∼1mM from 0 to 4mM and a liner response was observed from 0 to 0.2mM with a detection limit of ∼20μM. Compared with other methods, the carboxyl-modified silica nanoparticles (∼60nm) prepared by the reverse microemulsion method can serve as a stable and sensitive sensor platform because of their smaller size and facile conjugation with amine-containing molecules. In addition, the high sensitivity and selectivity of hoechst33258 was employed for the ssDNA and dsDNA determination, which takes advantage of the label-free aptamer and lower cost.
在这项工作中，我们开发了一种使用硅纳米粒子（NPs）作为平台和 Hoechst33258 作为信号报告物的简单灵敏的 ATP 检测方法。ATP 结合适体与固定在 NPs 上的探针 DNA（DNA(p)）杂交，在 NPs 表面形成适体/DNA(p)双链体。由于 ATP 结合适体在存在 ATP 的情况下从适体/DNA(p)双链体转变为适体/靶复合物，适体的构象变化导致 NPs 上的适体/DNA(p)双链体减少。通过分离硅纳米粒子并添加嵌入适体/DNA(p)（dsDNA）的 Hoechst33258，可以轻松实现 ATP 检测。已经证明了 ATP 和 CTP、GTP 或 UTP 之间具有良好的选择性，这是由于 ATP 适体与 ATP 之间的特异性识别。从 0 到 4mM 估算出 K(d)约为 1mM，从 0 到 0.2mM 观察到线性响应，检测限约为 20μM。与其他方法相比，通过反相微乳液法制备的羧基修饰的硅纳米粒子（约 60nm）由于其较小的尺寸和与含胺分子的简便缀合，可以作为稳定灵敏的传感器平台。此外，Hoechst33258 的高灵敏度和选择性被用于 ssDNA 和 dsDNA 的测定，这利用了无标记适体和低成本的优势。
