【摘要】：1,5-脫水葡萄糖醇(l,5-anhydroglueitol,1,5-AG)是一種反應(yīng)血糖水平的重要生物標(biāo)志物,臨床上常通過檢測人體血清中1,5-AG的濃度進行糖尿病的診斷、發(fā)展程度判斷、治療效果評估。論文瞄準(zhǔn)人體血清中1,5-AG的檢測問題,在LAPS芯片表面修飾吡喃糖氧化酶(Pyranose Oxidase,PROD)作為電化學(xué)探針,基于酶催化銀沉積原理,設(shè)計一種能特異性檢測1,5-AG的光尋址電位型電化學(xué)生物傳感器(Light Addressable Potentiometric Sensor,LAPS)。論文主要研究內(nèi)容如下:構(gòu)建LAPS生物傳感界面。以1,5-AG為測試對象,通過層層自組裝將能特異性識別1,5-AG的吡喃糖氧化酶修飾在LAPS芯片表面,構(gòu)建LAPS生物傳感界面;優(yōu)化PROD的固定化條件,通過在硅烷化的LAPS芯片上修飾納米金吸附層提高識別分子的負(fù)載量。利用納米金良好的生物相溶性和吸附作用將能特異性識別1,5-AG的PROD固定到LAPS芯片表面,增加被固定酶的穩(wěn)定性和有效保持被固定酶的生物活性。搭建基于Lab VIEW平臺的LAPS檢測系統(tǒng)。以ICL8038信號發(fā)生器芯片制作LAPS尋址光源的可調(diào)制驅(qū)動信號源,設(shè)計自動功率控制電路(Automatic Power Control:APC)控制尋址光源的驅(qū)動電流,使尋址光源輸出功率在工作溫度范圍內(nèi)保持恒定,利用Lab VIEW軟件搭建采集系統(tǒng)對數(shù)據(jù)進行采集、處理并繪制電流-電壓(I-V)曲線。基于酶催化銀沉積原理來實現(xiàn)對1,5-AG的特異性檢測。利用PROD對1,5-AG的催化作用,生成還原性物質(zhì)雙氧水。雙氧水將電解質(zhì)溶液中的銀離子還原為銀單質(zhì)使溶液中的銀離子濃度減小,導(dǎo)致LAPS芯片表面的膜電位發(fā)生變化,膜電位的變化也反映了1,5-AG濃度的變化。研究結(jié)果表明:電壓偏移量與1,5-AG濃度在60μg/m L至225μg/mL范圍內(nèi)具有良好的線性關(guān)系,相關(guān)系數(shù)為0.9977,檢測限為40μg/m L,相對標(biāo)準(zhǔn)偏差為1.37%,1,5-AG的回收率在92.95-103.50%范圍內(nèi),并且特異性良好。
[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.
【學(xué)位授予單位】：桂林電子科技大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：TP212

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