Infectious Research Center, Korea Research Institute of Bioscience & Biotechnology (KRIBB), Daejeon, 34141, Republic of Korea.
Department of Neuroscience, Graduate School, Kyung Hee University, Seoul, 02447, Republic of Korea.
Sci Rep. 2020 Feb 28;10(1):3772. doi: 10.1038/s41598-020-60715-x.
In this study, ultrasensitive and precise detection of a representative brain hormone, dopamine (DA), was demonstrated using functional conducting polymer nanotubes modified with aptamers. A high-performance aptasensor was composed of interdigitated microelectrodes (IMEs), carboxylated polypyrrole nanotubes (CPNTs) and DA-specific aptamers. The biosensors were constructed by sequential conjugation of CPNTs and aptamer molecules on the IMEs, and the substrate was integrated into a liquid-ion gating system surrounded by pH 7.4 buffer as an electrolyte. To confirm DA exocytosis based on aptasensors, DA sensitivity and selectivity were monitored using liquid-ion gated field-effect transistors (FETs). The minimum detection level (MDL; 100 pM) of the aptasensors was determined, and their MDL was optimized by controlling the diameter of the CPNTs owing to their different capacities for aptamer introduction. The MDL of CPNT aptasensors is sufficient for discriminating between healthy and unhealthy individuals because the total DA concentration in the blood of normal person is generally determined to be ca. 0.5 to 6.2 ng/mL (3.9 to 40.5 nM) by high-performance liquid chromatography (HPLC) (this information was obtained from a guidebook "Evidence-Based Medicine 2018 SCL " which was published by Seoul Clinical Laboratory). The CPNTs with the smaller diameters (CPNT2: ca. 120 nm) showed 100 times higher sensitivity and selectivity than the wider CPNTs (CPNT1: ca. 200 nm). Moreover, the aptasensors based on CPNTs had excellent DA discrimination in the presence of various neurotransmitters. Based on the excellent sensing properties of these aptasensors, the DA levels of exogeneous DA samples that were prepared from PC12 cells by a DA release assay were successfully measured by DA kits, and the aptasensor sensing properties were compared to those of standard DA reagents. Finally, the real-time response values to the various exogeneous DA release levels were similar to those of a standard DA aptasensor. Therefore, CPNT-based aptasensors provide efficient and rapid DA screening for neuron-mediated genetic diseases such as Parkinson's disease.
在这项研究中，使用适配体修饰的功能导电聚合物纳米管实现了对代表性脑激素多巴胺 (DA) 的超灵敏和精确检测。高性能适配体传感器由叉指微电极 (IME)、羧基化聚吡咯纳米管 (CPNT) 和 DA 特异性适配体组成。生物传感器通过 CPNT 和适配体分子在 IME 上的顺序共轭构建，基底集成到由 pH 7.4 缓冲液包围的液体离子门控系统中作为电解质。为了基于适配体传感器确认 DA 的胞吐作用，使用液体离子门控场效应晶体管 (FET) 监测 DA 的灵敏度和选择性。通过控制 CPNT 的直径来优化适配体传感器的最小检测限 (MDL；100 pM)，因为它们对适配体引入的不同能力导致其 MDL 不同。CPNT 适配体传感器的 MDL 足以区分健康个体和不健康个体，因为通过高效液相色谱法 (HPLC) 确定正常人血液中的总 DA 浓度通常约为 0.5 至 6.2 ng/mL (3.9 至 40.5 nM)（该信息是从由首尔临床实验室出版的“2018 年循证医学指南”中获得的）。与较宽的 CPNTs (CPNT1：约 200nm) 相比，直径较小的 CPNTs (CPNT2：约 120nm) 表现出 100 倍的更高灵敏度和选择性。此外，基于 CPNTs 的适配体传感器在存在各种神经递质的情况下具有出色的 DA 区分能力。基于这些适配体传感器的出色传感性能，成功地通过 DA 试剂盒测量了由 PC12 细胞通过 DA 释放测定制备的外源性 DA 样品的 DA 水平，并将适配体传感器的传感性能与标准 DA 试剂进行了比较。最后，对各种外源性 DA 释放水平的实时响应值与标准 DA 适配体传感器的响应值相似。因此，基于 CPNT 的适配体传感器为帕金森病等神经元介导的遗传疾病提供了有效的快速 DA 筛选。
