發(fā)布時(shí)間：2018-04-29 01:17

  本文選題：毛細(xì)管電泳 + 前沿分析法　； 參考：《中國(guó)人民解放軍軍事醫(yī)學(xué)科學(xué)院》2010年碩士論文

【摘要】： 高靈敏蛋白質(zhì)檢測(cè)方法及相互作用研究對(duì)疾病的診治具有重要意義,同時(shí)也是蛋白質(zhì)組學(xué)得以迅速發(fā)展的巨大動(dòng)力。近年來,針對(duì)功能蛋白的高靈敏度分析檢測(cè)新方法研究,一直是生命科學(xué)研究中蛋白質(zhì)分析領(lǐng)域的研究熱點(diǎn)。本論文即開展了基于毛細(xì)管電泳(CE)技術(shù)的相關(guān)蛋白質(zhì)間接檢測(cè)新方法及與小分子藥物或單鏈DNA適配體的結(jié)合作用研究,全文共分為四章。 第一章綜述了蛋白質(zhì)的高靈敏度檢測(cè)與識(shí)別技術(shù)概況,重點(diǎn)總結(jié)了近年來CE技術(shù)在蛋白質(zhì)分離分析、相互作用中的相關(guān)進(jìn)展。 第二章中,我們首次將具有高效分離能力的CE方法和BCA法結(jié)合起來,建立了一種間接檢測(cè)蛋白質(zhì)的新方法。以β-環(huán)糊精為包合添加劑,有效地實(shí)現(xiàn)了BCA-Cu+復(fù)合物和過量游離BCA分子的分離,通過微波輔助反應(yīng)達(dá)到BCA-Cu+快速、完全顯色的目的,實(shí)現(xiàn)了在低波長(zhǎng)處對(duì)轉(zhuǎn)鐵蛋白、蓖麻毒素等功能蛋白的快速、高靈敏度檢測(cè)的目的。本方法亦成功應(yīng)用于第一屆蓖麻毒素國(guó)際實(shí)驗(yàn)室間比對(duì)測(cè)試(2009年)蓖麻毒素蛋白樣品的檢測(cè)中。所測(cè)定的樣品中蓖麻毒素的含量結(jié)果與真值相比,偏差介于0.40-6.8%之間,說明了我們?cè)贑E中以BCA-Cu+復(fù)合物濃度的定量間接指代蛋白質(zhì)定量之思路的正確性。 第三章通過CE-前沿分析法(CE-FA)研究堿性藥物CBZ與HSA間相互作用,注重選擇R值處于較窄范圍的不同濃度的CBZ-HSA體系進(jìn)行測(cè)定,從而使線性方程與非線性方程所得到的結(jié)合參數(shù)符合較好,方便地求出了該體系的結(jié)合常數(shù)和結(jié)合比,該弱相互作用體系的解離常數(shù)(Kd)值為7.09×10-5M,CBZ與HSA近似為1:1結(jié)合,CBZ結(jié)合到蛋白上的結(jié)合百分?jǐn)?shù)介于15.8-22.4%之間。 第四章使用CE-FA法,結(jié)合激光誘導(dǎo)熒光檢測(cè)和適配體標(biāo)記技術(shù),以多種方式(R-Cf非線性擬合、平臺(tái)峰高變化-濃度間的非線性擬合、單點(diǎn)測(cè)定法)深入探討和表征了凝血酶適配體F-29mer和凝血酶間的結(jié)合作用,并詳細(xì)討論了三種方法所各適用的范圍。
[Abstract]:The study of highly sensitive protein detection and interaction is of great significance to the diagnosis and treatment of diseases, and it is also a huge driving force for the rapid development of proteomics. In recent years, high sensitivity analysis and detection of functional proteins has been a hot topic in the field of protein analysis in life sciences. In this paper, a new method for indirect detection of related proteins based on capillary electrophoresis (CE) technique and its binding to small molecular drugs or single-stranded DNA aptamers were studied. The whole thesis is divided into four chapters. In the first chapter, the high sensitivity detection and recognition technology of protein is reviewed, and the progress of CE in protein separation and interaction is emphatically summarized. In the second chapter, we combine CE method with BCA method for the first time, and establish a new method for the indirect detection of protein. Using 尾 -cyclodextrin as inclusion additive, the separation of BCA-Cu complex and excessive free BCA molecules was realized effectively. The aim of rapid and complete color development of BCA-Cu was achieved by microwave-assisted reaction, and transferrin at low wavelength was realized. The aim of rapid and high sensitivity detection of ricin and other functional proteins. This method has also been successfully applied to the determination of ricin samples in the first International Laboratory comparison Test of ricin (2009). Compared with the true value, the deviation of the determination of ricin content in the sample is between 0.40-6.8%, which indicates the correctness of our idea of quantifying protein indirectly with the concentration of BCA-Cu complex in CE. In chapter 3, the interaction between CBZ and HSA was studied by CE-frontier analysis method, and the determination of CBZ-HSA with different concentrations in a narrow range of R values was emphasized. Thus, the binding parameters obtained by linear equation and nonlinear equation are in good agreement, and the binding constant and binding ratio of the system are obtained conveniently. The dissociation constant (Kd) of the weak interaction system is 7.09 脳 10 ~ (-5) M ~ (-1) C _ (Z) and HSA is approximately 1:1 bound to protein. The percentage of binding to protein is between 15.8-22.4%. In chapter 4, we use CE-FA method, combined with laser-induced fluorescence detection and aptamer labeling, to fit the R-Cf nonlinearity in many ways, and the nonlinear fitting between peak height of platform and concentration. The binding between thrombin aptamer F-29mer and thrombin was investigated and characterized, and the suitable ranges of the three methods were discussed in detail.
【學(xué)位授予單位】：中國(guó)人民解放軍軍事醫(yī)學(xué)科學(xué)院
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2010
【分類號(hào)】：R341

【參考文獻(xiàn)】

相關(guān)期刊論文 前5條

1 趙燕燕,楊更亮,范子琳,王德先,陳義;高效毛細(xì)管電泳前沿分析法研究酸性藥物那格列奈與人血漿白蛋白的結(jié)合[J];分析化學(xué);2003年07期

2 閆妍;張淑珍;唐吉軍;顧明松;馮建林;謝劍煒;;一種基于肽質(zhì)量指紋譜技術(shù)鑒定蓖麻毒素的新方法[J];分析化學(xué);2006年02期

3 鄭麗輝;鄭新宇;童萍;陳國(guó)南;張?zhí)m;;毛細(xì)管電泳電化學(xué)淌度移動(dòng)法測(cè)定利尿劑和牛血清白蛋白結(jié)合常數(shù)[J];分析化學(xué);2009年01期

4 王忠慧;王惠民;王躍國(guó);;毛細(xì)管電泳在脂蛋白分析中的應(yīng)用[J];醫(yī)學(xué)檢驗(yàn)與臨床;2007年02期

5 單瑞峰,劉道杰;毛細(xì)管電泳在測(cè)定理化參數(shù)中的應(yīng)用[J];聊城大學(xué)學(xué)報(bào)(自然科學(xué)版);2005年01期

，

本文編號(hào)：1817741