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論文名稱 Title |
電噴灑輔助雷射脫附游離法的開發以及在蛋白質分析與分子影像上的應用 Development and Applications of Electrospray-Assisted Laser Desorption Ionization for Protein Analysis and Molecular maging |
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系所名稱 Department |
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畢業學年期 Year, semester |
語文別 Language |
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學位類別 Degree |
頁數 Number of pages |
237 |
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研究生 Author |
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指導教授 Advisor |
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召集委員 Convenor |
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口試委員 Advisory Committee |
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口試日期 Date of Exam |
2008-01-18 |
繳交日期 Date of Submission |
2008-02-15 |
關鍵字 Keywords |
雷射脫附、分子影像、固體、大氣壓力、電噴灑輔助雷射脫附游離法 Ambient, Solid, Laser Desorption, molecular imaging, ELDI / MS |
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統計 Statistics |
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中文摘要 |
電噴灑輔助雷射脫附游離質譜法(Electrospray-assisted Laser Desorption Ionization Mass Spectrometry, ELDI/MS)是一個結合電噴灑和雷射脫附的新游離技術,其特點是分析樣品不需前處理或添加基質,在一大氣壓力環境快速對固體樣品中所含的化學組份進行偵測。其游離過程是以雷射轟擊固體分析物表面,脫附出的中性的氣態分子再與經由電噴灑所產生的帶電液滴反應融合,經由電噴灑後游離過程產生分析物離子訊號。研究中介紹ELDI/MS原理、游離機制,並探討各個影響訊號的實驗參數以及後續改良進行探討。在固體大分子的應用主要分析各式生物體液(如血液、眼淚、唾液及血清)及組織(如肝臟、心臟及腦部)內蛋白質組成,並應用在醫療診斷,做為疾病的快速篩選技術以及蛋白體學的鑑定工具。在固體小分子的應用主要是強調快速鑑定固體樣品中所含的化學組份,包含藥片、顏料、高分子聚合物等,可應用在材料分析、食品安全及刑事鑑定上。最後是結合影像質譜分析法(Imaging Mass Spectrometry)直接對固態樣品進行表面掃瞄分析,研究中以牛樟芝及靈芝做為分析物,完成蕈類組織分子影像,及對植物組織進行活體(in vivo)分析。 |
Abstract |
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目次 Table of Contents |
目錄 論文摘要..................................................................................................Ⅰ 目錄..........................................................................................................Ⅱ 圖目錄…………………………………………………...……………...Ⅶ 壹、 緒論.................................................................................................1 一、 質譜學概述……………………………………………….………1 二、 質譜游離源………………………………………………….……3 三、 真空質譜法………………………………………………….……6 四、 大氣壓力質譜法…………………………………………….…...12 五、 大氣壓力固態游離質譜法……………………………………....24 六、 影像質譜法…………………………………………………...….37 七、 論文目標…………………………………….…………….……..43 貳、實驗部分..........................................................................................44 一、電噴灑輔助雷射脫附游離質譜法裝置...........................................44 (一)質譜儀部分...............................................................................45 1.離子阱質譜儀...........................................................................45 2.四極-飛行時間質譜儀…………………………………..……45 3. 三段式四極柱質譜儀…………………….……….……….46 4.基質輔助雷射脫附飛行時間質譜儀…………….………46 (二)電噴灑游離源部分...............................................................46 1.熔融系毛細管………………………………...…………..46 2.注射式幫浦.........................................................................47 3.高電壓電源供應器..............................................................47 4.質譜入口不鏽鋼管..............................................................47 (三)雷射脫附游離源部分..............................................................47 1. 脈衝式紫外光氮氣雷射.....................................................47 2. 脈衝式Nd-YAG紅外線雷射…………………………….48 (四)XYZ移動平台...........................................................................48 1. XYZ平台..............................................................................48 2. XY自動掃瞄平台.................................................................48 (五)CCD攝影機..............................................................................48 二、影像處理軟體.................................................................................49 三、離子模擬軟體..................................................................................49 四、化學藥品與製備..............................................................................49 參、ELDI / MS發展、原理、游離過程與機制………………………51 一、電噴灑輔助雷射脫附游離質譜法(ELDI / MS)發展………….51 二、ELDI / MS原理、游離過程與機制………………………………63 (一)ELDI / MS原理…………………………………………..63 (二)ELDI / MS游離過程…………………………..…………64 (三)ELDI / MS游離機制………………………….………….66 肆、ELDI / MS游離源界面設計改良與參數探討……….…………..69 一、ELDI / MS游離界面的改良……………………………………...69 二、ELDI / MS實驗參數探討………………………….……………..71 (一)雷射脫附與電噴灑游離法影響.........................................72 (二)雷射脫附影響 .................................................... ..............75 (三)MS入口不鏽鋼管………………..……………………….76 (四)ESI高電壓…………………………………………………77 (五)ESI毛細管內徑…………………………………………….81 (六)ESI毛細管至質譜入口距離……………………………….83 (七)ESI溶液流速……………………………………………….85 (八)ESI乾燥氣體………………………………………………..88 (九)ESI霧化氣體……………………………………………….91 (十)雷射角度………………………………………..…………93 (十一)雷射能量…………………………………………………95 (十二)雷射與ESI平行方向照射位置…………………………..97 (十三)雷射與ESI垂直方向照射位置……………….……………99 (十四)樣品盤與ESI毛細管距離……………………...................101 (十五)樣品盤材質………………………….…..……………….104 (十六)不同次雷射脫附………………….………………………105 (十七)雷射頻率………………………………………..….……..107 (十八)樣品盤移動速度…………………….…..………………..108 (十九)電噴灑溶液組成…………………………………………..110 (二十)不同質譜儀連接………………………………………….111 (二十一)基質影響……………………………………………113 (二十二)ELDI / MS偵測靈敏度 ……………………………118 伍、ELDI / MS應用於生物體內大分子蛋白質的分析……………..122 一、混和蛋白質標準品分析………………………………….……..122 二、以ELDI / MS直接分析牛奶內含組成……………………………124 三、以ELDI / MS直接分析全血內蛋白質組成………………..……126 四、以ELDI / MS直接分析人類眼淚、唾液及血清內蛋白質...........128 五、以ELDI / MS直接分析腦部中灰質及白質內部組成....................129 六、以ELDI / MS直接分析豬肝及豬心組織內含蛋白質部組成........135 陸、ELDI / MS直接偵測各式固體內含小分子的分析………………143 一、以ELDI / MS直接分析食用染料…………….………………….143 二、以ELDI / MS直接分析高分子聚合物……………………………145 三、以ELDI / MS直接分析彩色顏料……………….……………….146 四、以ELDI / MS直接分析市售光碟…………………………….….150 五、以ELDI / MS直接分析各式藥錠………………………….…….152 六、以IR-ELDI / MS直接分析各種食品……………………………..155 七、以IR-ELDI / MS直接分析活體(In Vivo)植物組織……………158 柒、ELDI / MS應用於質譜影像的分析……………………………..160 一、ELDI影像質譜分析法的概念…………………………………….160 二、ELDI影像質譜分析法的開法…………………………………..162 三、以ELDI Imaging進行植物組織分子影像的開發……………….168 (一)靈芝………………………………………………………..168 (二)牛樟芝……………………………………………………..173 捌、結論………………………………………………………………176 玖、參考資料………………………………………………………….178 |
參考文獻 References |
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