Development of highly integrated enzyme switches as a multi-parameter sensing device

酶开关是一种基于聚苯胺（PAn）膜电导测量的新型生物传感装置。与传统的酶传感器相比，酶开关可能具有以下优点：1)传感芯片本身具有记忆功能；2)响应可能与电极尺寸无关；3)结构简单，易于小型化和集成化。4)易于与执行机构连接，因为电阻变化是指标。在本研究中，通过使用制备的酶开关验证了这些优点。聚丙烯腈因氧化而失去电导率，并通过还原恢复电导率。如果用PAn制备氧化酶-过氧化物酶（HRP）共固定化电极，当生成的化合物I和II在PAn表面还原时，PAn可能被氧化，失去导电性。通过监测PAn的电导率，可以构建“酶开关”，通过特定底物的存在发出“关闭”信号。在酸性条件下，在两个微图纹薄膜电极上电化学制备PAn薄膜。然后用戊二醛交联法将葡萄糖氧化酶（或乳酸氧化酶）和HRP固定在PAn膜上。通过添加底物，可以观察到酶反应导致PAn电导率下降。在0.1 V vs.Ag/AgCl的作用下，氧化后的PAn恢复了电导率。在流动系统中，从“开”到“关”或“关”到“开”大约需要3 min，峰高与底物浓度相对应。所制备的酶开关具有酶试纸和酶传感器的双重优点。在酶反应过程中不需要施加电压，反应可以保持至少1小时。在多通道电极上构建了葡萄糖和乳酸的集成多通道酶开关。集成酶开关对葡萄糖或乳酸的反应是独立的，没有观察到化学串扰。

Enzyme switch is a novel biosensing device based on the conductometric measurements of polyaniline(PAn) film covered with enzymes. The enzyme switch might have following advantages over conventional enzyme sensors : 1)Sensing chip itself has a memory function, 2)response might be independent from the electrode size, 3)Miniaturization and integration are easy because of its simple structure. 4)Easy connection with actuators, since the resistance change is index. In this study, these advantages were verified by using prepared enzyme switches.PAn loses its electroconductivity by oxidation and recovers it by reduction. If the oxidase-peroxidase(HRP) co-immobilized electrodes are prepared with PAn, when the produced compounds I and II are reduced at the surface of PAn,PAn might be oxidized and may lose its conductivity. By monitoring conductivity of PAn, "enzyme-switch", which gives "off" signal by the existence of specific substrate, might be constructed.PAn film was prepared electrochemically over the two micro-patterned thin film electrodes under the acidic conditions. Then glucose oxidase(or lactate oxidase) and HRP were immobilized onto the PAn film by glutaraldehyde crosslinking method. By the addition of substrates, the decrease of PAn conductivity due to the enzyme reaction could be observed. The oxidized PAn recovered its conductivity by applying 0.1 V vs.Ag/AgCl. In flow system, it took approximately 3 min for switching from "on" to "off", or "off" to "on", and peak heights were correspond to substrate concentration. The prepared enzyme switch had advantages both as enzyme test strips and enzyme sensors. Voltage application was not necessary during the enzyme reaction, and the response could be preserved for at least 1h.An integrated multi-channel enzyme switch for glucose and lactate was constructed onto the multi-channel electrodes. The integrated enzyme switch showed responses to glucose or lactate independently and the chemical cross-talk was not observed.