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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
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dc.contributor.advisor | 何國榮 | |
dc.contributor.author | Chu-Yun Tsai | en |
dc.contributor.author | 蔡竺芸 | zh_TW |
dc.date.accessioned | 2021-06-13T16:48:22Z | - |
dc.date.available | 2005-07-04 | |
dc.date.copyright | 2005-07-04 | |
dc.date.issued | 2005 | |
dc.date.submitted | 2005-06-27 | |
dc.identifier.citation | 1. Wilkins, M. R., Sanchez, J. C., Golley, A. A., Appel, R. D.,
HumpherySmith, I., Hochstrasser, D. F., Williams, K. L. Genet. Eng. Rev. 1996, 13, 19 2. Wilkins, M. R., Sanchez, J. C., Williams, K. L., Hochstrasser, D. F. Electrophoresis 1996, 17, 830 3. Cleveland, D.W., et al. J Biol Chem, 1977, 252, 1102 4. Klose, J., U. Kobalz Electrophoresis, 1995, 16, 1034 5. Zeleny, J. Phy. Rev. 1917, 10, 1 6. Yamashita, M. and Fenn, J. B. J. Phys. Chem. 1984, 88, 4451 7. Yamashita, M. and Fenn, J. B. J. Phys. Chem. 1984, 88, 4671 8. Fenn, J. B., Mann, M., Meng, C. K., Wong, S. F., and Whitehouse, C. M. Science, 1989, 246, 64 9. M Dole, R. L. Hines, L. L. Mack, R. C. Mobley, L. D. Ferguson and M. B. Alice J. Chem. Phys. 1968, 49, 2240 10.Iribarne, J. V. and Thomson, B. A. J. Phys. Chem. 1976, 64, 2287 11.Licklider, L.J. Thoreen, C.C. Peng, J. Gygi, S.P. Anal. Chem. 2002, 74, 3076 12. Hookeun Lee, Eugene C. Yi, Bo Wen, Timothy P. Reily, Lance Pohl,Sidney Nelson, Ruedi Aebersold, David R. Goodlett, J. Chromatogr. B. 2004, 803, 101 13. Kelly, J. F.; Ramaley, L.; Thibault, P. Anal. Chem. 1997, 69, 51 14. Valaskovic, G. A.; McLafferty, F. W. Rapid Commum. Mass Spectrom. 1996, 10, 825 15. Chang YZ, Her GR. Anal. Chem. 2000, 72, 626. 16. Fong WY, Chan TW. J. Am. Soc. Mass Spectrom. 1999, 10, 72 17. Jianjun Zhai, Wenqing Shui, Xuejiao Xu, Yanling Yu, Huimin Bao, Xian Chen, Pengyuan Yang Rapid Commun. Mass Spectrom. 2004, 18, 1177 | |
dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/38834 | - |
dc.description.abstract | 我們以簡單的設備建立毛細液相層析電灑法串聯質譜儀,成功地鑑定大白鼠第二型肺泡細胞特有胞器-層質體 (lamellar bodies) 之組成蛋白質,有助於未來臨床上對呼吸窘迫症病患的處理與治療。有鑑於層析方法過於耗時,本研究第二部分為開發簡單且可拋棄式的灑探針,將消化並淨化後的胜肽直接送入串聯質譜儀進行分析,可達到快速鑑定蛋白質及無樣品殘留之問題;以肌紅蛋白 (Myoglobin) 的胰蛋白酶 (trypsin) 消化胜肽為樣品,蛋白質身分鑑定的極限可至2.5 fmol,具有使用方便及低樣品消耗的優點。第三部分我們以濃縮為主訴求,希望能開發簡單、實用且能線上淨化的方法,所開發之線上淨化微電灑串聯質譜儀,可大幅提高濃縮倍率及減少分析時間,實際應用於層質體的蛋白質體,得到高可信度的蛋白質鑑定結果。 | zh_TW |
dc.description.abstract | We set up an on-line micro capillary reversed phase liquid chromatography interfaced to a tandem mass spectrometry (μLC-MS/MS) for protein identification. Proteins from lamellar bodies which were separated by two-dimensional gel electrophoresis were identified by μLC-MS/MS. However, the current technique is rather time consuming. To reduce the analysis time, we developed a disposable device that combined on-line clean up and infusion unseparated proteolytic peptides into an ESI tandem MS. It was demonstrated that several proteins of lamellar bodies could be successfully identified by using this on-line clean up method. Compared to μLC-MS/MS, this method could identify proteins rapidly and without the problem of cross-contamination although the peptide sequence coverage was slightly lower. | en |
dc.description.provenance | Made available in DSpace on 2021-06-13T16:48:22Z (GMT). No. of bitstreams: 1 ntu-94-R92223058-1.pdf: 2205449 bytes, checksum: 9d30615723236426aa71ac587259050a (MD5) Previous issue date: 2005 | en |
dc.description.tableofcontents | 第一章 序論 ……………………………………………………………………… 1
1-1 蛋白質身分之鑑定 ………………………………………………… 3 1-2 二維膠體電泳 (2-Dimentional Polyacrylamide Gel Electrophoresis) …… 5 1-3 電灑法串聯質譜儀 (ESI Tandem MS) …………………………… 7 1-3-1 電灑法 (electrospray ionization;ESI) ……………………………… 7 1-3-2 離子阱質譜儀 ……………………………………………………… 9 第二章 實驗 …………………………………………………………………… 18 2-1 毛細液相層析系統 ……………………………………………………… 18 2-1-1 毛細管柱的製作 …………………………………………………… 18 2-1-2 流速與分流器 (Splitter) 的設計 ………………………………… 20 2-1-3 動相與梯度系統 …………………………………………………… 20 2-2 拋棄式微電灑探針 (Microelectrospray Tip) …………………………… 22 2-2-1 微電灑探針的製作 ………………………………………………… 22 2-2-2 取樣方式 …………………………………………………………… 22 2-2-3 導電方式 …………………………………………………………… 23 2-3 線上淨化 (Clean-up) 微電灑 (Microelectrospray) 偵測 …………… 24 2-3-1 淨化毛細管柱 (Clean-up column) ……………………………… 24 2-3-2 微電灑探針的製作 ………………………………………………… 25 2-4 電灑法串聯質譜儀 ……………………………………………………… 26 2-5 資料庫搜尋 ……………………………………………………………… 27 2-6 層質體 (Lamellar Bodies) 樣品製備 ………………………………… 28 2-6-1 蛋白質的定量 ……………………………………………………… 28 2-6-2 樣品去鹽處理 ……………………………………………………… 29 2-6-3 二維膠體電泳 (Two-Dimensional Electrophoresis;2-DE) ………… 30 2-6-4 膠體染色 (Staining) ………………………………………………… 32 2-6-5 膠體內水解 (In gel digestion) ……………………………………… 33 2-6-6 ZipTipC18淨化 (Clean-up) ………………………………………… 35 第三章 結果與討論 …………………………………………………………… 51 3-1 毛細液相層析電灑法串聯質譜儀 ……………………………………… 51 3-1-1 毛細液相層析系統設計…………………………………………… 53 3-1-2 毛細液相層析串聯質譜儀系統測試 ……………………………… 54 3-1-3 層質體 (Lamellar Bodies) 蛋白質身分鑑定 …………………… 54 3-2 拋棄式微電灑探針串聯質譜儀 ………………………………………… 56 3-2-1 微電灑探針訊號強度與穩定性 …………………………………… 57 3-2-2 拋棄式微電灑探針串聯質譜儀於蛋白質身分鑑定 ……………… 57 3-3 線上淨化 (Clean-up) 微電灑 (Microelectrospray) 偵測 …………… 59 3-3-1 線上淨化 (Clean-up) 微電灑 (Microelectrospray) 方法可行性 … 59 3-3-2 蛋白質鑑定之測試 ………………………………………………… 60 3-3-3 線上淨化 (Clean-up) 微電灑探針與拋棄式微電灑探針之比較 …………………………………………………………………… 61 3-3-4 線上淨化微電灑串聯質譜儀於層質體之蛋白質身分鑑定 ……… 61 第四章 結論 …………………………………………………………………… 83 參考文獻 ………………………………………………………………………… 84 | |
dc.language.iso | zh-TW | |
dc.title | 毛細液相層析與拋棄式微電灑探針電灑法質譜/質譜於電白質鑑定之開發與應用 | zh_TW |
dc.type | Thesis | |
dc.date.schoolyear | 93-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 林萬寅,傅明仁 | |
dc.subject.keyword | 質譜,蛋白質, | zh_TW |
dc.subject.keyword | mass spectrometry,protein identification, | en |
dc.relation.page | 85 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2005-06-27 | |
dc.contributor.author-college | 理學院 | zh_TW |
dc.contributor.author-dept | 化學研究所 | zh_TW |
顯示於系所單位: | 化學系 |
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