Kwon Seong Jung, Yang Haesik, Jo Kyungmin, Kwak Juhyoun
Department of Chemistry, Korea Advanced Institute of Science and Technology, Daejeon, Korea.
Analyst. 2008 Nov;133(11):1599-604. doi: 10.1039/b806302h. Epub 2008 Aug 11.
Redox cycling of enzymatically amplified electroactive species has been widely employed for high signal amplification in electrochemical biosensors. However, gold (Au) electrodes are not generally suitable for redox cycling using a reducing (or oxidizing) agent because of the high background current caused by the redox reaction of the agent at highly electrocatalytic Au electrodes. Here we report a new redox cycling scheme, using nicotinamide adenine dinucleotide (NADH), which can be applied to Au electrodes. Importantly, p-aminophenol (AP) redox cycling by NADH is achieved in the absence of diaphorase enzyme. The Au electrodes are modified with a mixed self-assembled monolayer of mercaptododecanoic acid and mercaptoundecanol, and a partially ferrocenyl-tethered dendrimer layer. The self-assembled monolayer of long thiol molecules significantly decreases the background current of the modified Au electrodes, and the ferrocene modification facilitates easy oxidation of AP. The low amount of ferrocene on the Au electrodes minimizes ferrocene-mediated oxidation of NADH. In sandwich-type electrochemical immunosensors for mouse immunoglobulin G (IgG), an alkaline phosphatase label converts p-aminophenylphosphate (APP) into electroactive AP. The amplified AP is oxidized to p-quinoneimine (QI) by electrochemically generated ferrocenium ion. NADH reduces QI back to AP, which can be re-oxidized. This redox cycling enables a low detection limit for mouse IgG (1 pg mL(-1)) to be obtained.
酶促放大电活性物质的氧化还原循环已被广泛应用于电化学生物传感器的高信号放大。然而，由于在高电催化金电极上试剂的氧化还原反应会导致高背景电流，金（Au）电极通常不适合使用还原剂（或氧化剂）进行氧化还原循环。在此，我们报告了一种新的氧化还原循环方案，使用烟酰胺腺嘌呤二核苷酸（NADH），该方案可应用于金电极。重要的是，在没有黄递酶的情况下实现了由NADH介导的对氨基苯酚（AP）氧化还原循环。金电极用巯基十二烷酸和巯基十一醇的混合自组装单层以及部分二茂铁连接的树枝状聚合物层进行修饰。长硫醇分子的自组装单层显著降低了修饰金电极的背景电流，并且二茂铁修饰促进了AP的容易氧化。金电极上少量的二茂铁使二茂铁介导的NADH氧化最小化。在用于小鼠免疫球蛋白G（IgG）的夹心型电化学免疫传感器中，碱性磷酸酶标记将对氨基苯磷酸（APP）转化为电活性AP。放大后的AP被电化学生成的二茂铁离子氧化为对醌亚胺（QI）。NADH将QI还原回AP，AP可再次被氧化。这种氧化还原循环使得能够获得小鼠IgG的低检测限（1 pg mL(-¹)）。
