高等学校化学学报

高等学校化学学报

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利用互补核酸杂交富集金胶实现信号扩增的电化学凝血酶蛋白生物传感器研究

郑静1,2, 冯婉娟1, 程圭芳1, 黄翠华1, 林莉1, 何品刚1, 方禹之1   

    1. 华东师范大学化学系, 上海 200062;
    2. 上海工程技术大学化学化工学院, 上海 201620
  • 收稿日期:2007-04-12 修回日期:1900-01-01 出版日期:2007-12-10 发布日期:2007-12-10
  • 通讯作者: 何品刚

Approaching Signal Amplification of Electrochemical Biosensor for Thrombin Based on Enrichment of Gold Nanoparticles Through Hybridization with Complementary Oligonucleotide

ZHENG Jing1,2, FENG Wan-Juan1, CHENG Gui-Fang1, HUANG Cui-Hua1, LIN Li1, HE Pin-Gang1* , FANG Yu-Zhi1
  

    1. Department of Chemistry, East China Normal University, Shanghai 200062, China;
    2. Department of Chemistry and Chemical Engineering, Shanghai University of Engineering Science, Shanghai 201620, China
  • Received:2007-04-12 Revised:1900-01-01 Online:2007-12-10 Published:2007-12-10
  • Contact: HE Pin-Gang

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被引次数

23

摘要： 介绍了一种利用互补核酸杂交富集金胶实现信号扩增的蛋白质生物传感器. 以凝血酶蛋白为研究对象, 利用凝血酶蛋白相对应的两段核酸适配体, 将适配体Ⅰ固定在磁性颗粒上, 用于特异性地捕获蛋白, 将适配体Ⅱ标记金胶作为检测信标. 由凝血酶蛋白和相对应的两段核酸适配体构建三明治结构的凝血酶蛋白生物传感器. 另外, 再通过信标金胶上过剩的核酸适配体链与另一段标记有金胶的互补核酸进一步杂交, 获得金胶的选择性聚集, 实现了信号扩增. 通过信号扩增, 使此传感器的灵敏度大大提高, 对凝血酶蛋白的检测下限可达到4.52×10-15 mol/L. 平行测定浓度为7.47×10-14 mol/L的凝血酶8次, 其RSD为3.0%. 该生物传感器对凝血酶蛋白有很好的特异性, 其它蛋白如溶菌酶和牛血清白蛋白的存在对于检测没有影响.

关键词: 信号扩增, 核酸适配体, 凝血酶蛋白, 金胶, 电化学生物传感器

Abstract: In this paper, a signal amplified electrochemical biosensor for thrombin based on the enrichment of the gold nanoparticles through the hybridization with the complememtary oligonucleotide was presented. Thrombin, as a typical protein model, has two combination sites with aptamers. In the experiment, aptamer Ⅰ was immobilized on magnetic nanoparticles for capturing thrombin, and aptamer Ⅱ was labeled with gold nanoparticle to offer an excellent electrochemical signal transduction. Through the specific recognition for thrombin, a sandwich format of magnetic nanoparticle/thrombin/gold nanoparticle was fabricated. And the signal amplification was further implemented by the enrichment of gold nanoparticles through the hybridization with the gold labeled complementary oligonucleotide，the resulting hybridization is capable of realizing more gold nanoparticle markers attaching to each target protein. A significant enhancement of the sensitivity was obtained for the detection of thrombin with the signal amplification. This new strategy allows the detection limit of the target protein down to 4.52×10-15 mol/L. The relative standard deviation of eight replicate determinations of 7.47×10-14 mol/L thrombin was 3.0%. The presence of other proteins such as BSA and lysozyme did not affect the detection of thrombin, which indicates that a high specificity of thrombin detection can be achieved.

Key words: Signal amplification, Nucleic acid aptamer, Thrombin, Gold nanoparticle, Electrochemical biosensor

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