联吡啶钌脂质体的合成及其在电化学发光免疫分析中的应用

发布时间：2018-07-26 08:28

【摘要】：近年来,对疾病相关标志物的灵敏检测引起了现代生物化学和生物医学研究的广泛兴趣。基于抗原与抗体之间特异性结合建立的免疫分析方法成为定量检测疾病标志物的最重要的分析方法之一,因为该方法中免疫反应分子之间高的特异结合性,能大大减小干扰。与放射免疫、荧光、化学发光等免疫分析法相比,电化学免疫分析方法有很多固有的优势,如成本低,灵敏度高,携带方便等,因此出现了大量各种各样的电化学免疫传感器。但是,在免疫传感器的发展过程中,人们发现抗原-抗体免疫复合物本身所形成的信号很弱,灵敏度有限,而临床中对早期超灵敏的生物标志物筛查癌症的需求不断增加。为此,出现了很多的信号放大策略。其中,将脂质体用于免疫传感器的信号放大引起了人们的极大兴趣,因为它具有制备简单,比表面积大,内部能包裹大量的信号分子等优点。本论文基于把电化学发光技术的优势和脂质体的特点结合起来,构建了电化学发光免疫传感器。为检验其性能,将其用于人免疫球蛋白G的检测,结果表明这种免疫传感器可以实现现代免疫传感器所要求的高灵敏度、高选择性、稳定及可再生等方面的研究目标。具体的研究工作包括以下几个方面:(1)以联吡啶钌为例,系统地研究了电化学发光脂质体的合成条件,建立了一种新的测定脂质体包封率的方法,另外还考察了脂质体的后处理如超声、挤压、渗析等对其包封率的影响;(2)基于碳纳米管修饰的玻碳电极以层层组装的方法,构建脂质体电化学发光免疫传感器,以抗体标记的吡啶钌脂质体为二抗,采用三明治型模式检测人IgG;(3)基于金纳米粒子和脂质体的信号放大作用,建立了脂质体电化学发光免疫传感器,实现了对人IgG双重信号放大的超灵敏检测,该传感器同时具有高灵敏度、高选择性、长期稳定性及易于再生等性能;(4)对联吡啶钌脂质体电化学发光免疫传感器在竞争检测模式中的应用进行了初步探讨,该检测模型可以实现对只有单一抗体的抗原的检测。本文为超灵敏免疫传感器的构建提供了一种新思路,在临床诊断中有潜在的应用价值。
[Abstract]:In recent years, sensitive detection of disease-related markers has attracted extensive interest in modern biochemistry and biomedical research. The immunoassay based on the specific binding between antigen and antibody has become one of the most important analytical methods for quantitative detection of disease markers, because of the high specificity of the immunoreactive molecules in this method, the interference can be greatly reduced. Compared with radioimmunoassay, fluorescence and chemiluminescence methods, electrochemical immunoassay has many inherent advantages, such as low cost, high sensitivity, easy to carry and so on. Therefore, a large number of electrochemical immunosensors have emerged. However, in the development of immunosensor, it has been found that the antigen-antibody immune complex itself is very weak and has limited sensitivity, and the demand for early super-sensitive biomarker screening for cancer is increasing in clinic. For this reason, there are a lot of signal amplification strategies. Among them, the signal amplification of liposome used in immunosensor has attracted great interest, because it has the advantages of simple preparation, large specific surface area, and the ability to encapsulate a large number of signal molecules inside. Based on the advantages of electroluminescent technology and the characteristics of liposomes, an electrochemiluminescence immunosensor was constructed in this paper. In order to test its performance, it has been applied to the detection of human immunoglobulin G. The results show that this immunosensor can achieve the research goals of high sensitivity, high selectivity, stability and regeneration required by modern immunosensors. The specific research work includes the following aspects: (1) taking the bipyridine ruthenium as an example, the synthesis conditions of electroluminescent liposomes were systematically studied, and a new method to determine the entrapment efficiency of liposomes was established. In addition, the effects of post-treatment of liposome such as ultrasonic, extrusion and dialysis on the entrapment efficiency were investigated. (2) Liposome electrochemiluminescence immunosensor was constructed by layer assembly method of glassy carbon electrode modified with carbon nanotubes. Using the antibody labeled pyridine ruthenium liposome as the second antibody, the sandwich pattern was used to detect human IgG. (3) based on the signal amplification of gold nanoparticles and liposomes, a liposome electrochemiluminescence immunosensor was established. The hypersensitivity detection of human IgG double signal amplification is realized. The sensor has high sensitivity and selectivity. (4) the application of electrochemiluminescence immunosensor of pyridine ruthenium liposome in competitive detection mode was discussed, and the detection model could be used to detect the antigen with only a single antibody. This article provides a new idea for the construction of hypersensitive immunosensor and has potential application value in clinical diagnosis.
【学位授予单位】：安徽师范大学
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：O657.3;R392.1

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