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研究生： 張國威
Chang, Kuo-Wei
論文名稱： 肺癌研究之微流體實驗室晶片
Microfluidic Labchip for Lung Cancer Studies
指導教授： 劉承賢
Liu, Cheng-Hsien
口試委員: 盧向成
Lu, Shiang-Cheng
陳致真
Chen, Chih-Chen
莊校奇
Chuang, Hsiao-Chi
李岡遠
Lee, Kang-Yun
學位類別： 博士
Doctor
系所名稱： 工學院 - 動力機械工程學系
Department of Power Mechanical Engineering
論文出版年： 2016
畢業學年度： 104
語文別： 英文
論文頁數： 87
中文關鍵詞： 肺癌細胞 、標靶治療 、濃度梯度 、介電泳 、細胞排列
外文關鍵詞： gradient
相關次數： 點閱：3  下載：0 
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    • 在世界各地，肺癌是癌症死亡的首要原因。大約有80-85 ％的肺癌是屬於非小細胞肺癌（NSCLC），在過去35年來，提高惡性腫瘤類型的生存率達五年的比例只有16 %。即使在早期非小細胞肺癌患者中，大約有一半會復發，儘管手術/放射線治療和輔助化療。因此，尋找更好的治療藥物增強對肺癌的活動仍在繼續。
    本研究設計一整合型標靶藥物檢測系統，透過系統中的電極設計將細胞以仿肺臟組織的形狀作排列，以進行體外的標靶藥物測試。此整合性標靶藥物檢測系統主要由產生濃度梯度的微流道設計和利用介電泳進行細胞排列的腔室所組成。當施予適當頻率的交流電壓時會產生介電泳,其依照細胞表面被極化的程度不同來操控細胞，以模仿肺組織之形態及增加細胞和細胞之間的交互作用。複雜的微流道結構能夠提供想要濃度梯度以同時地對肺癌細胞進行不同藥物劑量測試，並藉由量測體外藥物劑量的反應來預估實際體內肺癌細胞抑制效果。
    我們成功整合藥物檢測系統來評估艾瑞莎(Iressa)對肺癌細胞抑制測試，以驗證此系統的可行性。而且，個人化醫療檢驗已慢慢成為未來之趨勢，若能在體外仿照病人體內實際情況來進行檢測，更能夠貼近病人本身之檢驗，也能增加檢測的準確性。因此，在生物醫藥應用方面，整合型藥物檢測系統為一個很好的藥物及診斷工具。

    Lung cancer is still the leading cause of cancer deaths throughout the world. Approximately 80–85 % of all lung cancers are classified as non-small-cell lung cancer (NSCLC), an aggressive tumor type with a 5 year survival rate of 16 % that has improved to some extent over the last 35 years. Even in patients with early stage NSCLC, about half will relapse despite surgery/radiation and adjuvant chemotherapy. Therefore, the search for better therapeutic agents with enhanced activity against lung Carcinoma cancer continues. This study presents an integrated drug screening system based on the lung mimetic tissue pattern for in vitro screening of targeted therapy. The integrated targeted therapy drug screening system constitutes of continuous concentration gradient platform and dielectrophoretic (DEP) cell patterning chambers. DEP forces are based on polarity difference to manipulate and pattern the heterogeneous cells for mimicking the inherent lung morphology and increasing the cell-cell interaction. The multiplexed microchannels, i.e. concentration gradient generator, allow simultaneous administration of multiple drug doses to functional lung cancer cells enabling the in vitro dose-dependent drug responses.
    The current trend in personalized healthcare shows that medical care is becoming as individualized as possible. The goal is to not only provide customized diagnostic tests but also targeted medicines and dosages more precisely and safely to each patient. The integrated drug screening platform can serve as an effective tool for biomedical applications and diagnostics.

    摘要 I ABSTRACT II Acknowledgements III List of Figures VIII Chapter 1 Introduction - 1 - 1.1 Background and motivation - 1 - 1.2 Lab on a chip - 2 - 1.3 Tissue engineering - 3 - 1.3.1 Dielectrophoresis patterning to reconstruct tissue - 4 - 1.3.2 Cell patterning in micro-engineering gel - 5 - 1.3.3 Re-establish lung function on lab-on-a-chip - 6 - 1.3.4 Microfluidic flow field for 3D spheroid - 8 - 1.4 Cell culture in microfluidic system - 9 - 1.5 Gradient generator - 10 - 1.6 Active cell patterning - 14 - 1.6.1 Cell patterning using dielectrophoresis - 14 - 1.6.2 Cell patterning using negative dielectrophoresis - 14 - 1.6.3 Cell patterning using positive dielectrophoresis - 16 - 1.7 Lung cancer and targeted therapy - 19 - Chapter 2 Microsystem development for integrated targeted therapy - 20 - 2.1 Design principles - 20 - 2.1.1 Dielectrophoresis - 20 - 2.1.2 Transmembrane potential - 24 - 2.1.3 Mass transport on microscale - 24 - 2.2 Chip design concept - 26 - 2.2.1 Lithography of microchannel and electrode - 26 - 2.2.2 Microfluidic system gradient generator - 27 - 2.2.3 Hexagonal-shape electrode - 29 - 2.3 Numerical simulation - 30 - 2.3.1 Numerical simulation of gradient - 31 - 2.3.2 Numerical simulation of hexagonal-shape electrodes - 33 - 2.4 Microfabrication - 35 - 2.4.1 The microfabrication process - 35 - 2.4.2 Microfabrication results - 38 - Chapter 3 Materials and methods - 39 - 3.1 Chip design and system setup - 39 - 3.2 Biological experiments - 40 - 3.2.1 Cell culture medium preparation - 40 - 3.2.2 Low-conductive DEP buffer preparation - 41 - 3.2.3 Cell culture and preparation - 42 - 3.2.4 Operation parameters of DEP patterning processes - 44 - 3.2.5 Cell preparation for DEP manipulation - 46 - 3.2.6 Surface pretreatment for cell adhesion enhancement - 46 - 3.2.7 The parameter setup for DEP operation - 46 - 3.2.8 Instrument setup - 47 - 3.2.9 Cell patterning process - 48 - 3.3 Anticancer drug-Iressa - 50 - 3.4 MTT assay - 51 - 3.5 Statistical analysis - 53 - Chapter 4 Experiment results and discussion - 54 - 4.1 The results of gradient experiments - 54 - 4.2 The lung cancer tissue pattern on metal electrode - 56 - 4.2.1 Voltage and frequency of DEP operation parameter - 56 - 4.2.2 Cell patterning with positive/negative DEP - 57 - 4.2.3 HCC827/3T3 cells patterning - 59 - 4.2.4 Different ratio area of cell patterning - 60 - 4.3 MTT assay results - 61 - Chapter 5 Conclusions and future work - 69 - 5.1 Conclusions - 69 - 5.2 Future work - 71 - Reference - 73 -

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