血清髓过氧化物酶电流型免疫传感器的研究  

Amperometic Immunosensor for Myeloperoxidase in Serum

作　　者：周觅[1] 谢国明[1] 邹超世[1] 马霄[1] 熊兴良[2] 罗鹏[1] 刘成桂[3] 郑军[1] 

机构地区：[1]重庆医科大学检验系,临床检验诊断学教育部重点实验室,重庆400016 [2]重庆医科大学生物医学工程研究室,重庆400016 [3]四川省医学科学院附属医院检验科,简阳641400

出　　处：《分析化学》2009年第4期500-504,共5页Chinese Journal of Analytical Chemistry

基　　金：教育部临床检验诊断学重点实验室开放课题基金(No.200806)资助项目

摘　　要：将Nafion吸附到玻碳电极表面,并通过静电吸附和共价键合作用将硫堇和纳米金层层自组装到Nafion修饰的电极表面,然后通过形成的纳米金单层吸附髓过氧化物酶(MPO)抗体,最后用辣根过氧化物酶(HRP)封闭电极上的非特异吸附位点,同时起到放大响应电流信号的作用,研制了一种检测MPO的新型电流型免疫传感器。实验结果表明,该免疫传感器对MPO的响应特性良好,其线性检测范围为2.5～100μg/L;检出限为1.425μg/L,达到95%稳态响应时间<30s,批间、批内的平均RSD(n=20)<2.94%和4.15%。电极的稳定性良好,在连续30d内进行10次测量后,响应电流开始下降,平均测量值为初始的85.6%。探讨了抗体浓度、底物浓度、pH、温度及其它干扰物质等对该传感器的影响。使用本方法和经典的酶联免疫吸附实验(ELISA)同时对40份人血清标本MPO进行测试,结果表明:两者相关性良好(r=0.9971,p<0.0001)。该电流型免疫传感器具有灵敏度高、特异性好、不需标记和可以重复测量等优点。A novel amperometic immunosensor for the determination of myeloperoxidase （MPO） in human serum was developed by modifying Nation film on the glassy carbon electrode （GCE） , then thionine and colloidal gold were immobilized by static electricity and covalent bonding self-assembly. And then the antibody of MPO （anti-MPO） was absorbed on the surface of nano-gold monolayer. Finally, Horseridesh peroxidase （HRP） blocked the non-specific adsorption site on the electrode, meanwhile, the signal of response current was amplified. Experiment results showed that the good linear relations between peak current and concentration of MPO were established in the range of 2.5 - 100 μg/L. The detection limit was 1.4 μg/L and the response time reached 95% of the steady-state current in less than 30 s. The RSD values（n =20） of intra- batch and inter-batch were less than 2.9% and 4.2% , respectively. The stability of electrode was excellent, after having been surveyed 10 times in 30 days, the average value of response current began declining to 85.6% of the initial measurement. The influence of this immunosensor was investigated in terms of concentration of antibody, concentration of substrate, pH, temperature and other interferents. Correlation analysis showed that there was significant correlation between the method of immunosensor and ELISA in 40 clinical samples（r =0. 9971, p 〈0. 0001 ）. The proposed immunosensor for MPO is used for estimating the risk of aardiovascular disease.

关 键 词：髓过氧化物酶 生物传感器 纳米金 硫堇 化学修饰电极 

分 类 号：TP212.3[自动化与计算机技术—检测技术与自动化装置]