State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Biomedical Engineering Center, Hunan University, Key Laboratory for Bio-Nanotechnology and Molecule Engineering of Hunan Province, Changsha 410082, PR China.
Biosens Bioelectron. 2010 May 15;25(9):2101-6. doi: 10.1016/j.bios.2010.02.007. Epub 2010 Feb 20.
A sensitive and selective ligase-based signal-on electrochemical sensing method for adenosine-5'-triphosphate (ATP) detection had been developed using molecular beacon (MB)-like DNA. In this method, the biotin-tagged MB-like DNA was self-assembled onto a gold electrode to form a stem-loop structure by means of facile gold-thiol chemistry, which resulted in blockage of electronic transmission. It was eT OFF state. In the presence of ATP, two nucleotide fragments which were complementary to the loop of the MB-like DNA could be ligated by the ATP-dependent T4 DNA ligase. Hybridization of the ligated DNA with the MB-like DNA induced a significant conformational change in this surface-confined DNA structure, which in turn released the biotin from the surface allowing free exchange of electrons with the electrode generating a measurable electrochemical signal (eT ON). The resulting change in electron transfer efficiency was readily measured by differential pulse voltammetry at target ATP concentrations as low as 0.05 nM and with linear response range from 0.1 to 1000 nM. Moreover, it was also able to discriminate ATP from its analogues. The proposed method had been successfully applied to the determination of ATP in the Escherichia coli O157:H7 extracts of water samples, and the linear response was found between the concentrations of 10(3) and 10(7) cfu/mL.
基于连接酶的灵敏和选择性信号开启电化学传感方法已经被开发用于检测腺苷 5'-三磷酸（ATP），该方法使用分子信标（MB）样 DNA。在该方法中，生物素标记的 MB 样 DNA 通过简单的金-硫醇化学自组装到金电极上，形成茎环结构，从而阻止电子传递。此时处于关闭状态。在存在 ATP 的情况下，两个与 MB 样 DNA 环互补的核苷酸片段可以由 ATP 依赖的 T4 DNA 连接酶连接。连接的 DNA 与 MB 样 DNA 的杂交导致表面受限 DNA 结构发生显著的构象变化，从而将生物素从表面释放出来，允许与电极自由交换电子，产生可测量的电化学信号（开启状态）。在目标 ATP 浓度低至 0.05 nM 时，通过差分脉冲伏安法很容易测量到电子转移效率的这种变化，并且线性响应范围从 0.1 到 1000 nM。此外，它还能够区分 ATP 与其类似物。该方法已成功应用于水样中大肠杆菌 O157:H7 提取物中 ATP 的测定，并且在 10(3)到 10(7) cfu/mL 的浓度范围内发现了线性响应。
