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研究生: 郭于寧
Kuo, Yu-Ning
論文名稱: 以矽奈米線微米圖像陣列導引大鼠嗜鉻性瘤細胞株神經突生長位置方向
Guiding the Neurite Growth Position and Direction of PC12 Cells on Silicon-nanowire Micropatterns
指導教授: 嚴大任
Yen, Ta-Jen
口試委員: 王子威
Wang, Tzu-Wei
陳嘉勻
Chen, Chia-Yun
學位類別: 碩士
Master
系所名稱: 工學院 - 奈米工程與微系統研究所
Institute of NanoEngineering and MicroSystems
論文出版年: 2011
畢業學年度: 99
語文別: 英文
論文頁數: 77
中文關鍵詞: 矽奈米線微米圖像大鼠嗜鉻性瘤細胞株神經突
外文關鍵詞: silicon nanowires, micropattern, PC12 Cell, Neurite
相關次數: 點閱:2下載:0
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  • In the last decade, silicon nanowires (SiNWs) have attracted more attentions due to their excellent electrical and optical properties, and they also promise great applications in diverse fields including Raman silicon laser, antireflective solar cell, and highly sensitive biological sensors. Recently, the application of SiNWs has been extended into the biological field based on their controllable electrical property and superhydrophilic property. In addition to their electrical and surface property, a great amount of studies indicated that nano-topography can enhance cellular viability and physiological functions. Therefore, SiNWs may play a significant role as an in vitro cell culture matrix due to the aforementioned properties.
    For in vivo cell culture, it should be noticed that a neuron is an electrically excitable cell that transmits information by electrochemical impulses. Unfortunately, the function of the mammalian nervous system still remains unclear. Therefore, some researchers attempt to culture neurons in vitro to operate fundamental studies or to reverse-engineer the nervous system. However, neurons grow and differentiate into randomly connected neuronal populations on commercial culture plates. Researchers can merely measure behaviors of homogeneous neuronal circuits. The response of structured organization and controlled connectivity similar to those found in the nervous system are lacking. Consequently, we propose to figure out the cytotoxicity of mouse fibroblasts (L929) and pheochromocytom neurons (PC12) on various types of SiNWs and flat Si chips, for analyzing their cellular metabolic activity, cell adhesion and growth behaviors. Furthermore we also attempt to culture neurons on SiNWs chips with micropatterns in order to guide neurites growth direction for in vitro neuronal engineering.
    In this study, we fabricated SiNWs with different lengths employing the EMD method. Our results indicate that SiNWs are well aligned on Si chips and the diameter falls between 103.7-248.2 nm. Such highly rough and well-aligned SiNWs therefore possess superhydrophilic properties that may favor cell adhesion and following growth behavior and differentiation. We found that SiNWs can be regarded as a non-cytotoxic matrix when cultured with L929 and PC12, and further provide a better affinity to enhance cell adhesion compared to flat Si and nitride layer. These phenomena dominantly depend on the superhydrophilic and highly rough surface of SiNWs enabling to enhance high protein adsorption rate during cell adhesion.
    By combining different cell-adhesion affinities of SiNWs and nitride onto a line-pattern substrate, the majority of neurons culturing on SiNW micropatterns with 5-μm width of line pattern selectively grow and extend along SiNWs patterns instead of nitride region. The selective growth direction supposed to be guided by the rough surface and superhydrophilic property of SiNWs allowing higher protein adsorption than nitride. The ability of SiNW-micropattern chips for guiding neurite successfully demonstrated in this study will be a potential tool for neuronal engineering in the future.


    Content I List of Figures IV List of Tables VIII 誌謝 IX Abstract X Chapter 1 Introduction 1 1.1 Silicon nanowires 1 1.2 Motivation 2 Chapter 2 Literature Review 3 2.1 The Neuron 3 2.1.1 Structure 3 2.1.2 Cytoskeleton 4 2.1.3 Focal adhesion 5 2.2 Fundamental studies in mammalian nerve systems 6 2.3 Cell patterning techniques 8 2.3.1 Photolithography 8 2.3.2 Soft lithography 9 2.4 Silicon nanowires 12 2.4.1 Approaches of the synthesis of SiNWs 12 2.4.2 Electroless Metal Deposition (EMD) Method 16 2.5 Wettability of materials 19 2.5.1 Young’s equation 20 2.5.2 Cassie-Baxter model 21 2.6 Cell model 22 Chapter 3 Experimental Procedure 24 3.1 Fabrication of SiNWs 24 3.1.1 EMD method for SiNWs fabrication 24 3.1.2 Diameter analysis 25 3.2 SiNW Micropatterns 26 3.2.1 Chip design 26 3.2.2 The protection mask 26 3.2.3 The steps of SiNW micropatterns fabrication process 26 3.3 Cell growth on SiNW micropatterns 30 3.3.1 Cell culture 30 3.4 Measurement and Analysis Methods 30 3.4.1 Contact angle measurement 30 3.4.2 Surface topography measurement 31 3.4.3 Living cell observation 31 3.4.4 Cell viability assay 32 3.4.5 Cell morphology observation 33 3.4.6 Immunofluorescence staining of cells 34 Chapter 4 Results and Discussion 36 4.1 The fabrication of SiNWs 36 4.1.1 SiNWs arrays via EMD method 36 4.1.2 Different lengths of SiNWs 38 4.2 Cell growth on SiNWs 41 4.2.1 L929 cell line 41 4.2.2 Contact angle measurement on different substrates 48 4.2.3 PC12 cell line 50 4.3 Guiding the neurite growth of PC12 cells on SiNW micropatterns 55 4.3.2 Morphology of SiNW micropatterns 56 4.3.3 Guiding the neurite growth of PC12 cells on 20-μm-length SiNW micropatterns 59 4.3.4 Guiding the neurite growth of PC12 cells on 1-μm-length SiNW micropatterns 64 Chapter 5 Conclusions 67 Reference 68

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