【摘要】：近年來,對疾病相關標志物的靈敏檢測引起了現(xiàn)代生物化學和生物醫(yī)學研究的廣泛興趣�；诳乖c抗體之間特異性結合建立的免疫分析方法成為定量檢測疾病標志物的最重要的分析方法之一,因為該方法中免疫反應分子之間高的特異結合性,能大大減小干擾。與放射免疫、熒光、化學發(fā)光等免疫分析法相比,電化學免疫分析方法有很多固有的優(yōu)勢,如成本低,靈敏度高,攜帶方便等,因此出現(xiàn)了大量各種各樣的電化學免疫傳感器。但是,在免疫傳感器的發(fā)展過程中,人們發(fā)現(xiàn)抗原-抗體免疫復合物本身所形成的信號很弱,靈敏度有限,而臨床中對早期超靈敏的生物標志物篩查癌癥的需求不斷增加。為此,出現(xiàn)了很多的信號放大策略。其中,將脂質體用于免疫傳感器的信號放大引起了人們的極大興趣,因為它具有制備簡單,比表面積大,內(nèi)部能包裹大量的信號分子等優(yōu)點。 本論文基于把電化學發(fā)光技術的優(yōu)勢和脂質體的特點結合起來,構建了電化學發(fā)光免疫傳感器。為檢驗其性能,將其用于人免疫球蛋白G的檢測,結果表明這種免疫傳感器可以實現(xiàn)現(xiàn)代免疫傳感器所要求的高靈敏度、高選擇性、穩(wěn)定及可再生等方面的研究目標。具體的研究工作包括以下幾個方面:(1)以聯(lián)吡啶釕為例,系統(tǒng)地研究了電化學發(fā)光脂質體的合成條件,建立了一種新的測定脂質體包封率的方法,另外還考察了脂質體的后處理如超聲、擠壓、滲析等對其包封率的影響;(2)基于碳納米管修飾的玻碳電極以層層組裝的方法,構建脂質體電化學發(fā)光免疫傳感器,以抗體標記的吡啶釕脂質體為二抗,采用三明治型模式檢測人IgG;(3)基于金納米粒子和脂質體的信號放大作用,建立了脂質體電化學發(fā)光免疫傳感器,實現(xiàn)了對人IgG雙重信號放大的超靈敏檢測,該傳感器同時具有高靈敏度、高選擇性、長期穩(wěn)定性及易于再生等性能;(4)對聯(lián)吡啶釕脂質體電化學發(fā)光免疫傳感器在競爭檢測模式中的應用進行了初步探討,該檢測模型可以實現(xiàn)對只有單一抗體的抗原的檢測。本文為超靈敏免疫傳感器的構建提供了一種新思路,在臨床診斷中有潛在的應用價值。
[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.
【學位授予單位】：安徽師范大學
【學位級別】：碩士
【學位授予年份】：2010
【分類號】：O657.3;R392.1

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