這篇論文研究了幾種基於聚二甲基硅氧烷（PDMS）的微流控芯片，及其對於酶反應和蛋白質結晶的應用。 / 本論文分為兩部份。首先，論文第一部份介紹了兩種微流控芯片。利用到PDMS透氣的性質，第一種微流控芯片是一種基於脫氣PDMS的納升液體進液系統。在第一種微流控芯片的基礎上，我們引進一種可由普通注射器控制的氣閥控制系統，可對反應液體的分配和混合進行更方便和精確的操作。兩種芯片均成功應用於酶反應動力學的測定。在一次實驗中，只需要3~5微升的反應物就可以得到鹼性磷酸酶（alkaline phosphatase）的Michaelis-Menten動力學。 / 論文第二部份研究了“體積效“應對於蛋白質結晶的影響。首先，基於微孔的微流控芯片的蛋白質結晶篩選實驗揭示了在小體積（納升）下，蛋白質的結晶條件比在大體積（微升）下更多。在液滴里的蛋白質結晶實驗結果說明大體積的蛋白質液滴結晶速度更快。最後，蛋白質結晶實驗成功在雙乳液（double emulsion）的內核中進行。 / This thesis describes the design and development of micro dispensing systems for enzyme assay and for protein crystallization. The micro dispensing systems were fabricated by the soft-lithography method with the widely used material poly(dimethylsiloxane) (PDMS), which is gas and water permeable elastomer. / This thesis contains two major parts. In the first part, enzyme assay was performed in two micro dispensing systems, one based on microwells and the other based on pneumatic valves. The complete Michaelis-Menten kinetics measurement of the alkaline phosphatase (AP) with different fluorescein diphosphate (FDP) was achieved in one chip for each system using the fluorescence detection. These micro dispensing systems’ fabrication and operation were simple, and the total sample consumption was about 3~5 μL. / The second part reports the study of the volume effect on protein crystallization. Three micro dispensing systems, the microwell-based, droplet-based and double emulsion-based systems, were developed to perform the protein crystallization. Lysozyme and thaumatin were chosen as the model proteins. First, the protein crystallization screening experiments showed that protein crystallized in more precipitant conditions in the microchip than in conventional microbatch system. Second, the protein crystallization results carried out in droplets showed that protein crystallized faster in larger droplets. Finally, the protein crystallization in double emulsions was demonstrated. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Zhou, Xiaohu. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 87-91). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese. / Abstract --- p.i / 摘要 --- p.iii / Acknowledgements --- p.iv / Table of Contents --- p.vi / List of figures --- p.viii / Chapter Chapter 1 --- Introduction to microfluidics --- p.1 / Chapter 1.1 --- Fabrication of microfluidic devices --- p.2 / Chapter 1.1.1 --- PDMS-based microfluidic device --- p.2 / Chapter 1.1.2 --- PMMA microfluidic device --- p.6 / Chapter 1.1.3 --- Assembly of the microfluidic device --- p.8 / Chapter 1.2 --- Degassed PDMS pumping method --- p.9 / Chapter 1.3 --- Droplet-based microfluidics --- p.13 / Chapter 1.4 --- Thesis organization --- p.16 / Chapter Chapter 2 --- Micro dispensing systems for enzyme assay --- p.17 / Chapter 2.1 --- Introduction to enzyme assay --- p.17 / Chapter 2.1.1 --- Introduction to micro platforms for enzyme assay --- p.17 / Chapter 2.1.2 --- Introduction to enzyme kinetics and Michaelis-Menten kinetics --- p.21 / Chapter 2.2 --- Experimental --- p.26 / Chapter 2.2.1 --- Design of the micro dispensing systems --- p.26 / Chapter 2.2.2 --- Fabrication of the microfluidic devices --- p.30 / Chapter 2.2.3 --- Reagents and operation --- p.32 / Chapter 2.3 --- Results and discussion --- p.32 / Chapter 2.3.1 --- Microwell-based dispensing system --- p.32 / Chapter 2.3.2 --- Micro dispensing system based on pneumatic valves --- p.40 / Chapter 2.4 --- Conclusions --- p.45 / Chapter Chapter 3 --- Micro dispensing systems for protein crystallization --- p.47 / Chapter 3.1 --- Introduction to protein crystallization --- p.47 / Chapter 3.1.1 --- Principle of protein crystallization --- p.49 / Chapter 3.1.2 --- From macrofluidics to microfluidics --- p.51 / Chapter 3.2 --- Experimental --- p.56 / Chapter 3.2.1 --- Design of the micro dispensing systems --- p.56 / Chapter 3.2.2 --- Fabrication of the microfluidic devices --- p.58 / Chapter 3.2.3 --- Reagents and operation --- p.60 / Chapter 3.2.4 --- Layer-by-layer modification --- p.61 / Chapter 3.3 --- Results and discussion --- p.65 / Chapter 3.3.1 --- Demonstration of the micro dispensing systems --- p.65 / Chapter 3.3.2 --- Protein crystallization screening results --- p.69 / Chapter 3.3.3 --- Protein crystallization in droplets --- p.73 / Chapter 3.3.4 --- Protein crystallization in w/o/w double emulsions --- p.77 / Chapter 3.4 --- Conclusions --- p.80 / Appendix --- p.82 / References --- p.87
| Identifer | oai:union.ndltd.org:cuhk.edu.hk/oai:cuhk-dr:cuhk_328025 |
| Date | January 2012 |
| Contributors | Zhou, Xiaohu., Chinese University of Hong Kong Graduate School. Division of Chemistry. |
| Source Sets | The Chinese University of Hong Kong |
| Language | English, Chinese |
| Detected Language | English |
| Type | Text, bibliography |
| Format | electronic resource, electronic resource, remote, 1 online resource (xi, 91 leaves) : ill. (some col.) |
| Rights | Use of this resource is governed by the terms and conditions of the Creative Commons “Attribution-NonCommercial-NoDerivatives 4.0 International” License (http://creativecommons.org/licenses/by-nc-nd/4.0/) |
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