簡易檢索 / 詳目顯示

回結果列表
研究生： 吳仁貴
Wu, Ren-Guen
論文名稱： 新型電化學式毛細管電泳微晶片之研發
A Novel Capillary Electrophoresis Microchip with Electrochemical Detection
指導教授： 張憲彰
Chang, Hsien-Chang
學位類別： 碩士
Master
系所名稱： 工學院 - 醫學工程研究所
Institute of Biomedical Engineering
論文出版年： 2003
畢業學年度： 91
語文別： 中文
論文頁數： 83
中文關鍵詞： 電化學檢測 、毛細管電泳晶片 、場效流控制 、自組性單層膜
外文關鍵詞： electrochemical detection, FEF-EOF control, capillary electrophoresis
相關次數： 點閱：114  下載：1 
分享至:
查詢本校圖書館目錄 查詢臺灣博碩士論文知識加值系統 勘誤回報

    • 現今對於分析微小之生物樣本，除了需要有高靈敏度的感測方法之外，更須搭配高解析效能之分離技術，故本研究利用毛細管電泳分離技術結合電化學檢測原理，以微機電製程方法製作微小化之電化學式毛細管電泳晶片，配合微流道幾何設計及局部場效控制電滲流之技術達到高靈敏度及高解析度之最佳化效能。實驗中所使用之電化學式毛細管電泳晶片是由玻璃電極晶片與毛細管電泳通道所組成。通道製作完成後，用氧電漿處理poly(dimethylsiloxane) PDMS晶片以增加表面之親水性，用來強化通道本身之電滲透流性質與接合強度。為了達到晶片於實驗之最佳化效能，對玻璃電極晶片與電泳通道做進一步的設計，在電泳通道方面，利用增加通道的寬度，縮小通道深度;在玻璃電極晶片方面，則改變電化學電極組的大小以增加電化學反應的面積和固定反應樣本的總體積。最後配合場效流控制(field effect flow control)之概念，利用自組性單層膜(self-assembled monolayer)修飾於金電極上當作絕緣層，此電極置於電泳通道的正下方，並外加一誘發電壓於此電極上用以極化此絕緣層進而控制微管道內之電滲流方向與大小。將此場效應控制電滲流之技術，配合外加電場提供一電驅動力來控制微流道內液體之流動，同時利用倒立式螢光顯微鏡之監測，來達成精確操控移動率為 μeof =7.235 x 104 cm2V-1s-1之場效應雙向電滲流，最後於晶片末端結合一電化學電極組。以此完成之新型電化學式毛細管電泳晶片可重複使用多達20次，此自主性單層膜的絕緣性範圍為 +0.8 V ~ -0.8 V，其崩潰電壓為正負1.0 V。同時應用於神經傳導物質(dopamine、catechol )之分離與檢測，此系統對dopamine之檢測濃度極限值可達6.25x10-8 M其S/N比為3.3，分離效率之理論板數為5847。本新型晶片之效能與一般電化學式電泳晶片之檢測濃度極限值(2~3x10-7 M)比較而言可降低5倍左右。

    We fabricated a three-electrode electrochemical detector and an electric decoupler on a same glass chip and integrated with an O2-plasma-treated PDMS micro-channel to form the electrochemical capillary electrophoresis (EC-CE) microchip. The platinized decoupler which is an electroplated platinum particle that could be usefully decouple the electrochemical detection circuit from the interference of an separation electric field in 10 mM 2-(Nmorpholino)ethanesulfonic acid (MES buffer, pH 6.0) solution. In order to improve the electrochemical property of gold reference electrodes, Pt pseudo- reference electrode was manufactured by depositing Pt particles on Au reference electrodes with 5 mM H2PtCl6 scanned from –500 mV to 500 mV at 100 mV/sec by cyclic voltammetry. The platinized pseudoreference electrode was demonstrated to offer a stable potential in electrochemical detection. In this EC-CE system, the limit of detection of dopamine was 0.125 μM at an S/N=3.8. The linear range of different concentrations for ideal responses of dopamine was found between 0.25 to 50 μM with a correlation coefficient of 0.9974 and a sensitivity of 11.76 pA/μM. In addition, a novel technique to control the electro-osmotic flow (EOF) has been fabricated by using the self-assembled monolayer (SAM) as an insulator of a field-effect flow control (FEF-EOF) system in our research. A perpendicular electric field of 2.7 MV/cm could be generated by an extra-induced gate voltage (Vg) of 0.8 V, at this condition, the electroosmotic mobility (μeof) ranges from -3.1 to 3.4 × 10-4 cm2/V•s by modulation of +0.8 V to -0.8 V regardless of buffer pH. The low power (< 1 V) consumption was due to the only 2.9 nanometers thickness SAM-insulator which contains 1-octadecanethiol (ODT) or the 11-mercaptoundecanoic acid (MUA). At the same time, we combined the electrochemical electrodes with FEF-EOF system to increase the resolution of the detection limitation of neurotransmitters. For dopamine, the theoretical plate numbers were increased from 2669 to 5847 by the electrochemical detection without or with FEF-EOF control. In this FEF-EOF system, the limit of detection of dopamine was 0.0625 μM at an S/N=3.3.

    中文摘要………………………………………………………….. I 英文摘要…………………………………………………………. II 誌謝……………………………………………………………….. III 目錄……………………………………………………………….IV 圖目錄……………………………………………………………VII 表目錄..…………….......………….…………………………........IX 符號單位...........................................................................................X 第一章 序論 1.1研究背景與目的...…………………………….......…………....1 1.2微實驗處理形生物晶片的發展...………………………...........3 1.2.1前處理型晶片……….....…………………………………3 1.2.2生化反應晶…………....………………………………….4 1.2.3檢測分析晶片………....………………………………….5 1.2.4國內外微實驗處理型晶片的應用與市場分析………....6 1.3電化學式毛細管電泳微晶片的發展與應用……………........10 1.3.1DNA基因晶片………….....…………………………….11 1.3.2蛋白質基因晶片...............................................................12 1.3.3免疫酵素晶片...................................................................13 1.3.4生物有機分子晶...............................................................14 1.4毛細管電泳原理........................................................................15 1.4.1電泳遷移率......................................................................15 1.4.2電滲流的原理..................................................................16 1.4.3毛細管電泳效能的評估..................................................16 1.5 毛細管電泳的效能評估……………………………………...18 1.5.1 理論板數與解析度.........................................................18 1.6毛細管電泳的偵測方法............................................................19 1.6.1紫外光可見光吸收偵測法...............................................19 1.6.2螢光偵測法.......................................................................20 1.6.3質譜偵測法.......................................................................20 1.6.4電化學偵測法...................................................................21 1.7 研究架構……………………………………………………...25 第二章 設備與方法........................................................................26 2.1 研究設備...................................................................................26 2.2 實驗藥劑與配製方法...............................................................27 2.3 微小化晶片製作.......................................................................28 2.3.1 PDMS通道晶片的製作................................................28 2.3.2 玻璃電極晶片的製作.....................................................29 2.3.3 鉑擬參考電極的製作.....................................................30 2.3.4 自組性單分子薄膜電極晶片的製作.............................30 2.4 實驗系統架構...........................................................................31 2.4.1 電化學式毛細管電泳系統架構.....................................31 2.4.2 場效應電場控制電滲流之系統架構.............................31 2.5 微機電製程技術.......................................................................32 2.5.1 金屬層真空蒸鍍............................................................32 2.5.2 光微影技術....................................................................33 2.6 實驗設計與方法.......................................................................34 2.6.1 實驗設計.........................................................................34 2.6.2 實驗方法.........................................................................35 2.6.2.1 毛細管電泳系統操控.............................................35 2.6.2.2 誘發場效電場之操控.............................................35 第三章 結果與結論........................................................................36 3.1毛細管電泳晶片的實驗結果................................................... 36 3.1.1鉑擬參考電極的性能驗證...............................................36 3.1.2毛細管電泳分離條件的探討...........................................37 3.1.3電泳晶片的樣本測試......................................................38 3.1.4再現性之評估...................................................................39 3.2場效應控制電滲流之實驗結果................................................40 3.2.1電滲流方向之控制..........................................................40 3.2.2電滲流流速之控制..........................................................41 3.2.3自組性單分子薄膜之絕緣性..........................................41 3.2.4自組性單分子薄膜之耐受性..........................................42 3.2.5雙向電滲流控制之證……..............................................42 3.3新型電化學式毛細管電泳晶片之實驗結果.........................43 3.3.1晶片幾何設計結合場效應電滲流測試..........................44 3.3.2檢測極限值之驗證..........................................................45 第四章 結論 4.1 毛細管電泳分離與電化學檢測之最佳實驗條件……...……46 4.2 場效應控制電滲流之最佳實驗條件…………………...……47 4.3 未來發展………………………………………………...……47 4.3.1 效應控制電滲流之流動波面曲線驗證………………47 4.3.2 雙T型通道晶片結合場效應控制電滲流之發展……48 第五章 參考文獻…………………………………………………49 自述............................................................................................83

    1. R. B. M. Schasfoort, S. Schlautmann, J. Hendrikse, A. van den Berg, “Field-Effect flow control for microfabricated fluidic networks”, Science, 199, 286, 942-945.
    2. C. C. Wu, R. G. Wu, J. G. Huang, H. C. Chang, “Three-electrode electrochemical detector and platinum film decoupler integrated with a capillary electrophoresis microchip for amperometric detection”, Analytical Chemistry, 2003, 75, 947-952.
    3. M. K. Jessamine, N. I. Gitlin, A. D. Stroock, “Components for integrated poly(dimethylsiloxane) microfluidic system”, Electrophoresis, 2002, 23, 3461-3473.
    4. D. J. Satake, N. S. Sekhon, J. Borninski, B. Rubinsky, “Instantaneous, quantitative single-cell viability assessment by electrical evaluation of cell membrane integrity with microfabricated devices”, Sensors and Actuators A, 2003, 105, 31-39.
    5. P. R. C. Gascoyne, J. Noshari, F. F. Becker, R. Pethig, “Use of dielectrophoretic collection spectra for characterizing difference between normal and cancerous cells”, IEEE Transactions on Industry Applications ,
    6. P. Y. Huang, X. B. Wang, J. P. H. Burt, “Positive and negative dielectrophoretic collection of colloidal particles using interdigitated castellated microelectrodes”, Journal of Physics D: Applied Physics, 1992, 24, 881.
    7. T. Fujii, “PDMS-based microfluidic devices for biomedical applications”, Microelectronic Engineering, 2002, 61–62, 907–914.
    8. A. T. Woolley, D. Hadley, P. Landre, A. J. deMello, R. A. Mathies, M. A. Northrup, “Functional integration of PCR amplification and capillary electrophoresis in a microfabricated DNA analysis device”, Analytical Chemistry, 1996, 68, 4081-4086.
    9. R. Y. Zhang, H. K. Lee, S. F. Y. Li, “Conventional capillary electrophoresis in comparison with short-capillary electrophoresis and microfabricated glass chip capillary electrophoresis for the analysis of fluorescein isothiocyanate anti-human immunoglobulin G”, Journal of Chromatography A, 1997, 781, 287-293.
    10. A. T. Woolley, K. Q. Lao, A. N. Glazer, R. A. Mathies, “Capillary electrophoresis chips with integrated electrochemical detection”, Analytical Chemistry, 1998, 70, 684-688.
    11. M. A. Burns, C. H. Mastrangelo, T. S. Sammarco, F. P. Man, J. R. Webster, B. N. Johnson, B. Forester, D. Jones, Y. Fields, A. R. Kaiser, D. T. Burke, “Microfabricated structures for integrated DNA analysis”, Proceedings of the National Academy of Sciences of the United Stated of America, 1996, 93, 5556-5561.
    12. Y. Murakami, M. Suda, K. Yokayama, T. Takeuchi, E. Tamiya, I. Karube, “Micromachined enzyme reactor for FIA system”, Microchemical Journal, 1994, 49, 319-325.
    13. G. Ramsay,” DNA chips: state-of-the art”, Nature Biotechnology, 1998, 16, 40-44.
    14. M. Hashimoto, K. Tsukagoshi, R. Nakajima, K. Konda, A. Arai, “Microchip capillary electrophoresis using on-line chemiluminescence detection”, Journal of Chromatography A, 2000, 867, 271-279.
    15. M. S. Talary, J. P. H. Burt, P. Pethig, “Future trends in diagnosis using laboratory-on-a-chip technologies”, Microelectronic Engineering, 1998, 117, 191-203.
    16. http://www.biochipmaster.com/
    17. http://www.aclara.com/
    18. http://www.cepheid.com/index3.html
    19. http://www.caliper.com/
    20. 鄭郅言博士，”微流體生醫晶片的研發與應用”，第三屆 生物微奈米技術研討會，2003年4月,台南
    21. P. V. Pathak, H. J. Fsuer, S. Ekston,”Capillary electrophoresis microchips with thick-film amperometric detectors: separation and detection of phenolic compounds”, Analytica Chimica Acta, 1990, 416, 9-14.
    22. A. G. Ewing, M. Nishizawa, T. Matsue, I. Uchida, “Penicillin sensor based on a microarray electrode coated with pH-responsive polypyrrole”, Analytical Chemistry, 1991, 63, 2642-2644.
    23. M. Lunte, L. S. L. Loranelle, O. D. Richard,U. Bilitewski, D. J. Harrison, “Bi-enzymatic and capillary electrophoretic analysis of non-fluorescent compounds in microfluidic devices: Determination of xanthine”, Sensors and Actuators B, 1993, 52, 369-376.
    24. M. C. Cheng, H. J. Huang, “Application of nickel-microelectrode-incorporated end-column detector for capillary electrophoresis determination of alditols and alcohols”, Analytica chemiva acta, 1996, 341, 83-90.
    25. J. A. Manz, A. T. Wood, A. N. Glazer, “Capliiary electrophoresis chips with integrated electrochemical detection”, Analytical Chemistry, 1992, 64, 684-688.
    26. T. wooley, M. A. Strausbauch, A. F. R. Huhmer, N. Borson, S. R. Jurrens, J. Craighead, P. J. Ewttstein, B. Eckloff, B. Kline, J. P. Landers, “Infrared-mediated thermocycling for ultrafast polymerase chain reaction amplification of DNA”, Analytical Chemistry, 1998, 70, 4361-4368.
    27. D. J. Weiss, C. E. Lunte, S. A. Soper, S. M. Ford, Y. C. Xu, “Nanoliter-scale sample preparation methods directly coupled to polymethylmethqacrylate-based microchips and gel-filled capillary for the analysis of oligonucleotides”, Journal of Chromatography A, 2000, 854, 107-120.
    28. J. A. Lapso, W. Trabesinger, G. J. Schutz, H. J. Gruber, H. Schindler, T. Schmidt, “Detection of individual oligonucleotide paring by single-molecule microscopy”, Analytical Chemistry, 2002, 74, 279-283
    29. C. S. Effenhauser, B. Haab, R. A. Mathies, “Single-molecule detection of DNA separations in microfabricated capillary eelectrophoresis chips employing focused molecular streams”, Analytical Chemistry, 1999, 71, p. 5137-5154
    30. L. Zhang., H. F. Arlinghaus, M. N. Kwoka, K. B. Jacobson, “Analysis of biosensor chips for identification of nucleic acids”, Analytical Chemistry, 2000, 72, 3747-3753.
    31. J. M. Herrero-Martinez, C. L. Colyer, S. D. Mangru, D. J. Harrison, “Microchip-based capillary electrophoresis of human serum proteins”, Journal of Chromatography A, 2001, 785, 271-276.
    32. M. Chatrathi, J. Wang, F. Vinet , P. Chaton, Y. Fouillet, “Microarrays and microfluidic devices: miniaturized systems for biological analysis”, Analytical Chemistry, 2001, 73, 41–47.
    33. J. A. Gawron, P. C. Wang, D. L. DeVoe, C. S. Lee, “Integration of polymeric membranes with microfluildic networks for bioanalytical applications”, Electrophoresis, 2000, 21, 3857-3867.
    34. J. Wang, I. Moser, G. Jobst, G. A. Urban, “Biosensor arrays for simultaneous measurement of glucose, lactate, glutamate, and glutamine”, Biosensors & Bioelectronics, 2001, 17, 297-302.
    35. P. M. Chatrathi, K. K. Jain, “Applications of biochip and microarray systems in pharmacogenomics”, Pharmacogenomics, 2001, 4, 289-307.
    36. A. Narvaez, G. suarez, J. Z. Zhu, C. Y. Tian, W. Wu, “Fabrication and characterization of glucose sensors based on a microarray H2O2 electrode”, Biosensors and Bioelectronics, 2000, 15, 295-300.
    37. M. Bratten, P. G. Righetti, “Capillary electrophoresis in analytical biotechnology” CRC Press, 1997.
    38. A. G. Ewing, M. Nishizawa, T. Matsue, I. Uchida, “Fabrication of a pH-sensitive microarray electrode and applicability to biosensors”, Sensors and Actuators B, 2000, 21, 53-56.
    39. R. S. Martin, A. J. Gawron, S. M. Lunte, “Dual-electrode electrochemical detection for PDMS-fabricated capillary electrophoresis microchips”, Analytical Chemistry, 2000, 72, 3196-3202.
    40. J. Wang, M. P. Chatrathi, B. M. Tian, “Capillary electrophoresis microchips with amperometric detectors: separation and detection of phenolic compounds”, Analytica Chimica Acta, 2002, 73, 9-14.
    41. P. D. Grossman, J. C. Colburn, Capillary electrophoresis: theory and practice, Academic Press, 1992.
    42. F. Foret, L. Křivánková, P. Boček, Capillary zone electrophoresis, VCH, 1993.
    43. A. Weston, P. R. Brown, HPLC and CE: principles and practice, Academic Press, 1997.
    44. S. F. Y. Li, Capillary electrophoresis: principles, practice and applications, Elsevier, 1993.
    45. D. yuzhong, K. Ikuta, S. Maruo, Y. Fukaya, T. Fujisawa, “Biochemical IC chip toward cell free DNA protein synthesis”, Electrophoresis, 1998, 17, 131-136.
    46. S. Wallingford, A. G. Ewing, D. Beebe, M. Wheeler, “Microfluidic technology for amperometric detection assisted reproduction”, Theriogenology, 1987, 57, 125-135.
    47. B. Huang, R. S. Mirtain, L. M. V. Lerberghe, “Three-Dimensional Micro-Channel Fabrication in Polydimethylsiloxane Elastomer”, Journal of Microelectromechanical Systems, 2000, 9, 76-81.
    48. T. D. Lu, P. Cassidy, M. pumera, A. E. Feldman, “Capillary electrophoresis-electrochemistry microfluidic system for the determination of organic peroxides” Jounal of Chromatography B, 1995, 898, 657-681.
    49. S. Wallingford, A. G. Ewing, P. E. Michel, “Polymer microfluidic chips for electrochemical and biochemical analysis“, Electrophoresis, 2002, 23, 858-867.
    50. H. Y. Wang, R. S. Foote, S. C. Jacobson, J. H. Schneibel, J. M. Ramsey, “Low temperature bonding for microfabrication of electrochemical analysis devices”, Sensors and Actuators B, 1997, 45, 199-207.
    51. A. H. Ye, S. C. Baldwin, “Towards disposable lab on a chip: PDMS microchip electrophorsis devices with electrchemical detection”, Sensors and Actuators B, 2000, 21, 53-56.
    52. C. Zhong, B. M. Lunte, Tracy L. Pzxon “Electrophoresis in nonometer inner Diameter Capillaries with Electrochemical Detection”, Analytical Chemistry, 1999, 70, 3196-3202.
    53. T. Boegal, S. C. Baldwin, A. Partonz, “Development of lab-on-a-chip technology using electrochemical detector on column of microchannel”, Journal of Micromechanical and Microengeering, 1993, 8, 57–63.
    54. R. S. Mirtain, G. C. Gerhardt, Adsorption-Based electrochemical detection of electrochemical analytes for capillary electrophoresis”, Analytical Chemistry, 1994, 66, 908-915.
    55. C. S. Lee, W. C. Blanchard, C. T. Wu, “Direct control of the electroosmosis in CZE by using an external electric field”, Analytical Chemistry, 1990, 62, 1550-1552.

    下載圖示 校內：2004-07-17公開
    校外：2004-07-17公開
    QR CODE