I. Physikalisches Institut, Justus-Liebig-Universität Gießen, Heinrich-Buff-Ring 16, 35392 Gießen, Germany.
Institut für Veterinär-Physiologie und -Biochemie, Justus-Liebig-Universität Gießen, Frankfurter Straße 100, 35392 Gießen, Germany.
Biosens Bioelectron. 2016 Mar 15;77:1048-54. doi: 10.1016/j.bios.2015.10.076. Epub 2015 Oct 30.
The response characteristics of acetylcholinesterase-modified AlGaN/GaN solution-gate field-effect transistors (AcFETs) are quantitatively analyzed by means of a kinetic model. The characterization shows that the covalent enzyme immobilization process yields reproducible AcFET characteristics with a Michaelis constant KM of (122 ± 4) μM for the immobilized enzyme layer. The increase of KM by a factor of 2.4 during the first four measurement cycles is attributed to partial denaturation of the enzyme. The AcFETs were used to record the release of acetylcholine (ACh) by neuronal tissue cultivated on the gate area upon stimulation by rising the extracellular K(+) concentration. The neuronal tissue constituted of isolated myenteric neurons from four to 12 days old Wistar rats, or sections from the muscularis propria containing the myenteric plexus from adult rats. For both cases the AcFET response was demonstrated to be related to the activity of the immobilized acetylcholinesterase using the reversible acetylcholinesterase blocker donepezil. A concentration response curve of this blocking agent revealed a half maximal inhibitory concentration of 40 nM which is comparable to values measured by complementary in vitro methods.
通过动力学模型对乙酰胆碱酯酶修饰的 AlGaN/GaN 溶液栅场效应晶体管 (AcFET) 的响应特性进行了定量分析。表征结果表明，共价酶固定化过程产生了可重复的 AcFET 特性，固定化酶层的米氏常数 KM 为（122±4）μM。在最初的四个测量循环中，KM 增加了 2.4 倍，这归因于酶的部分变性。将 AcFET 用于记录在通过升高细胞外 K(+)浓度刺激栅极区域上培养的神经元组织释放乙酰胆碱 (ACh)。神经元组织由 4 至 12 天大的 Wistar 大鼠的分离肠神经元或含有肠丛的肌层切片组成。对于这两种情况，使用可逆乙酰胆碱酯酶抑制剂多奈哌齐证明 AcFET 响应与固定化乙酰胆碱酯酶的活性有关。该抑制剂的浓度反应曲线显示半最大抑制浓度为 40 nM，与互补的体外方法测量的值相当。
