基于酶催化银沉积原理的1,5-脱水葡萄糖醇电化学传感器研究

发布时间：2019-01-10 13:50

【摘要】：1,5-脱水葡萄糖醇(l,5-anhydroglueitol,1,5-AG)是一种反应血糖水平的重要生物标志物,临床上常通过检测人体血清中1,5-AG的浓度进行糖尿病的诊断、发展程度判断、治疗效果评估。论文瞄准人体血清中1,5-AG的检测问题,在LAPS芯片表面修饰吡喃糖氧化酶(Pyranose Oxidase,PROD)作为电化学探针,基于酶催化银沉积原理,设计一种能特异性检测1,5-AG的光寻址电位型电化学生物传感器(Light Addressable Potentiometric Sensor,LAPS)。论文主要研究内容如下:构建LAPS生物传感界面。以1,5-AG为测试对象,通过层层自组装将能特异性识别1,5-AG的吡喃糖氧化酶修饰在LAPS芯片表面,构建LAPS生物传感界面;优化PROD的固定化条件,通过在硅烷化的LAPS芯片上修饰纳米金吸附层提高识别分子的负载量。利用纳米金良好的生物相溶性和吸附作用将能特异性识别1,5-AG的PROD固定到LAPS芯片表面,增加被固定酶的稳定性和有效保持被固定酶的生物活性。搭建基于Lab VIEW平台的LAPS检测系统。以ICL8038信号发生器芯片制作LAPS寻址光源的可调制驱动信号源,设计自动功率控制电路(Automatic Power Control:APC)控制寻址光源的驱动电流,使寻址光源输出功率在工作温度范围内保持恒定,利用Lab VIEW软件搭建采集系统对数据进行采集、处理并绘制电流-电压(I-V)曲线。基于酶催化银沉积原理来实现对1,5-AG的特异性检测。利用PROD对1,5-AG的催化作用,生成还原性物质双氧水。双氧水将电解质溶液中的银离子还原为银单质使溶液中的银离子浓度减小,导致LAPS芯片表面的膜电位发生变化,膜电位的变化也反映了1,5-AG浓度的变化。研究结果表明:电压偏移量与1,5-AG浓度在60μg/m L至225μg/mL范围内具有良好的线性关系,相关系数为0.9977,检测限为40μg/m L,相对标准偏差为1.37%,1,5-AG的回收率在92.95-103.50%范围内,并且特异性良好。
[Abstract]:Ln 5-anhydrogluetol 1-AG is an important biomarker of blood glucose level. The clinical diagnosis of diabetes mellitus is often made by detecting the concentration of 1-5-AG in human serum and judging the development of diabetes mellitus. Evaluation of therapeutic effect. Aiming at the detection of 1H 5-AG in human serum, the surface modification of pyrrolidone oxidase (Pyranose Oxidase,PROD) on the surface of LAPS chip was used as an electrochemical probe based on the principle of enzymatic silver deposition. A photoaddressable potentiometric electrochemical biosensor (Light Addressable Potentiometric Sensor,LAPS) was designed. The main contents of this paper are as follows: construct LAPS biosensor interface. In order to construct the LAPS biosensor interface, the sucrose oxidase, which can specifically recognize 15-AG, was modified on the surface of LAPS chip by self-assembly. The immobilization conditions of PROD were optimized, and the loading of recognition molecules was increased by modifying the gold nanoparticles on silanized LAPS chips. By using the good biocompatibility and adsorption of gold nanoparticles, the PROD which can specifically recognize 1-5-AG can be immobilized on the surface of LAPS chip, which can increase the stability of the immobilized enzyme and effectively maintain the biological activity of the immobilized enzyme. Build LAPS detection system based on Lab VIEW platform. The modulable driving signal source of LAPS addressable light source is fabricated by using ICL8038 signal generator chip. The automatic power control circuit (Automatic Power Control:APC) is designed to control the driving current of the addressing light source, so that the output power of the addressable light source is kept constant in the working temperature range. Lab VIEW software is used to set up a collection system to collect data, process and draw current-voltage (I-V) curves. Based on the principle of enzymatic silver deposition, the specific detection of 1 ~ 5-AG was realized. Hydrogen peroxide was formed by the catalytic action of PROD on 1 ~ 5-AG. The reduction of silver ions in electrolyte solution to silver by hydrogen peroxide reduces the concentration of silver ions in the solution, which leads to the change of membrane potential on the surface of LAPS chip, and the change of membrane potential also reflects the change of 1-5-AG concentration. The results show that the voltage offset has a good linear relationship with the concentration of 1 渭 g / mL to 225 渭 g/mL. The correlation coefficient is 0.9977, the detection limit is 40 渭 g / mL, and the relative standard deviation is 1.37. The recoveries of 1C 5 AG were in the range of 92.95-103.50%, and the specificity was good.
【学位授予单位】：桂林电子科技大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：TP212

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