基于二氧化硅-硫堇纳米复合物构建高灵敏癌胚抗原电流型免疫传感器  

A highly sensitive amperometric immunosensor based on SiO_2-Thi nanocomposites for the detection of carcinoembryonic antigen

作　　者：蒋文[1] 田树高[1] 陈先玉[1] 程家蓉[1] 曲中堂[1] 

机构地区：[1]重庆医药高等专科学校,重庆医学检验试剂研究所,重庆401331

出　　处：《分析试验室》2017年第1期37-41,共5页Chinese Journal of Analysis Laboratory

基　　金：重庆市教委科学技术研究基金(KJ1502702);重庆市卫生计生委医学科研基金(20132151,2015MSXM097);重庆医药高等专科学校博士启动基金(YGZ2013301)资助

摘　　要：利用电沉积纳米金(AuNPs)修饰玻碳电极(GCE)表面并通过AuNPs固定癌胚抗原(CEA)的捕获抗体(Ab1),以牛血清白蛋白(BSA)封闭非特异性吸附位点;以γ-(2,3环氧丙氧)丙基三甲氧基硅烷(GPMS)作交联剂,将单分散的SiO_2纳米粒子与电子媒介体硫堇(Thi)结合成SiO_2-Thi纳米复合物,偶联辣根过氧化物酶(HRP)标记的CEA二抗(HRP-Ab2)作为电化学免疫检测信号,构建了具有信号放大效应的电流型免疫传感器并用于CEA的高灵敏检测。在CEA存在下,进行电化学酶联夹心免疫反应。在含有H2O2的磷酸盐缓冲溶液(PBS)中,标记在SiO_2-Thi纳米复合物上的HRP能催化H_2O_2氧化电子媒介体Thi,产生增强的还原峰电流,从而提高检测CEA的峰电流响应信号,进而实现对CEA的高灵敏电化学酶联夹心免疫分析。在最优实验条件下,该免疫传感器的差分脉冲伏安(DPV)还原峰电流与CEA质量浓度的对数在0.01~20ng/mL范围内呈良好的线性关系,检出限为3pg/mL(S/N=3)。该传感器对血清样品进行加标回收实验,回收率为97.3%~105.7%,可初步用于临床对CEA的检测。In this work, a novel amperometric immunosensor was developed by using thionine （Thi） functionalized silica nanocomposites （SiO2-Thi） to associate horseradish peroxidase （HRP）-labeled secondary antibody （HRP-Ab2 ） against carcinoembryonic antigen （CEA） as electrochemical signal amplification labels for the sensitive detection of CEA. Gold nanoparticles （AuNPs） were electrodeposited on a glassy carbon electrode （GCE） surface, and then the capture anti-CEA （AbI ） was anchored on the surface of AuNPs was used to block nonspecific by the adsorption of AuNPs and Abl. Bovine serum albumin （BSA） adsorption sites. SiO2-Thi nanocomposites were synthesized with monodispersed silica nanoparticles and thionine by using γ-glycidoxypropyhrimethoxysilane （GPMS） as the linkage and used to conjugate with HRP-Ab2 through EDC/NHS. Detection is based on the electrochemical active enzyme linked sandwich immunoassay. The amplified electrochemical signals derived from the carried SiO2-Thi nanocomposites toward the reduction of H202 using thionine as electron mediator and HRP as enhancer, and were measured by differential pulse vohammetry （DPV）. Under the optimal conditions, the DPV peak current was linearly dependent on the logarithm of CEA concentration in the range of 0. 01 - 20 ng/mL with a detection limit of 3.0 pg/mL （ S/N = 3 ）. Moreover, the immunosensor also displayed acceptable reproducibility, good stability, and excellent selectivity.Furthermore, the immunosensor was also used for the detection of CEA in spiked human serum samples with the recoveries of 97.3% - 105.7%, indicating its potential application in clinic detection of CEA.

关 键 词：电化学酶联夹心免疫分析 电流型免疫传感器 二氧化硅-硫堇纳米复合物 癌胚抗原 

分 类 号：O657.1[理学—分析化学]