基于电化学分子信标的多肿瘤标志物联检免疫芯片研究

项目名称： 基于电化学分子信标的多肿瘤标志物联检免疫芯片研究

项目编号： No.20805059

项目类型： 青年科学基金项目

立项/批准年度： 2009

项目学科： 金属学与金属工艺

项目作者： 戴宗

作者单位： 中山大学

项目金额： 20万元

中文摘要： 肿瘤标志物的灵敏和快速检测对肿瘤的早期发现、治疗，以及预后和复发的监测具有重要意义。本项目以建立肿瘤等重大疾病标志物的高灵敏检测技术为目的，基于双层磷脂膜和ITO电极，建立了抑制免疫分析中非特异性吸附的方法和超灵敏置换免疫分析技术；根据碱基在纳米材料修饰电极上的直接电催化氧化性质，结合重亚硫酸盐对甲基化胞嘧啶的特异性反应和限制性内切酶对特定序列的识别作用，以及DNA单碱基错配检测技术，在DNA整体甲基化水平，特异位点甲基化水平，以及p53肿瘤抑制基因甲基化状态的电化学检测方法取得新进展。同时，根据蛋白质在变性剂诱导下，结构变化引起生物活性和催化能力改变实现信号ON/OFF状态的控制，研究了自组装单层性质对血红蛋白直接电子传递行为的调控；探讨了氯离子、酸、尿素、盐酸胍等变性剂对血红蛋白去折叠的作用机制；进一步通过控制蛋白质结构，实现了对酶分子催化能力的调控和对富勒烯纳米材料的有效助溶，为设计电化学分子信标体系提供了新思路。本项目的研究将有助于肿瘤标志物的灵敏快速测定，为疾病的早期诊治提供新手段和依据。

中文关键词： 电化学分子信标；肿瘤标志物；DNA甲基化；免疫测定

英文摘要： Sensitive and rapid detection of tumor markers is of great significance for early cancer diagnosis. This project was aimed at developing effective and sensitive analytical approaches for tumor markers detection. Based on bilayer lipid membrane and ITO electrode, a novel dual-layer membrane cloaking method was proposed for overcoming the non-specific adsorption during immunoassay and an ultrasensitive displacement immunoassay method was raised. In view of the direct electrocatalytic oxidation of DNA bases, in combination of the specific bisulfite reaction and the restriction endonuclease reaction with one-mismatch analytical techniques, some progresses were obtained on electrochemical analysis of global DNA methylation, site-specific DNA methylation and the methylation state of p53 tumor suppressor gene. Besides, according to the fact that the ON/Off state of proteins signal can be manipulated through inducement of protein by denaturants, the effect of SAM property on the direct electron transfer of Hb was investigated, and the unfolding mechanisms of Hb by chloride, acid, urea and guanidine hydrochloride were studied. The mechanism was used to realize the manipulation of the catalytic ability of enzyme and the solubilization of fullerene, which will provide new approaches for construction of electrochemical molecular beacon. The investigations of the project will helpful for fast and sensitive detection of tumor markers, and providing new techniques for clinic diagnosis.

英文关键词： Electrochemical molecular beacon; tumor markers; DNA methylation; immunoassay