在生物科學的領域當中，探討細胞間的交互作用在生理機制的影響是一個很重要課題，也由於在生理系統上含有各種不同的細胞，不同種細胞的特徵與表現甚至是同種細胞間都不會完全相同，若給予外在的刺激，所釋放的訊號也會有所不同。為了要微觀的得取真實細胞的情況，現今已有各種探討單細胞間釋放物質的研究，而對於分析這些單細胞彼此間化學物質的傳遞也有許多種量測方式，但常見的困難點在於包含要先將複雜的樣本作預處理、濃縮，花時間且繁複，並由於單細胞經過刺激後在極短的時間內即會釋出化學分子，所以需要即時做好存取，並即時偵測，所以我們需要選擇一個整合性的平台，將各部分的工作有效率且有系統的結合在一個晶片上，以其能達到即時觀測這些化學分子隨著時間的變化。

此研究提出具兩大特色的微流道晶片，能調控電場自由選擇不同帶電性的樣本，並且能時序控制濃縮待測分子，將偵測極限降低，且不再需要再進行酵素或化學分子修飾，就能進行濃縮反應。本研究製作了一整合性的微系統平台，能夠將所有系統整合，期望能在每份樣本連續進入流道內時能夠及時做好濃縮，並且也能順序的做電化學訊號的偵測。濃縮效果是透過在奈米線區域給予電場加以極化，各奈米線間形成電雙層的帶電柵欄，將顆粒較大以及帶有相同電荷的分子濃縮在奈米線區域前，利用神經傳導物質具有特殊帶電性質，能作高效率的時序濃縮，透過濃縮將每段時間的訊號保存並放大，並將各時間收集的待測分子引領到電化學訊號之偵測區域，希望能觀察神經細胞的分泌物質隨著時間變化的結果，同時也能保有偵測極限低的特質。

致謝
摘要
目錄
表目錄
圖目錄
第一章 序論
1.1研究背景
1.2 研究動機
第二章 文獻回顧
2.1 導論
2.2 細胞培養與流體控制方法
2.3 濃縮方法
2.3.1微米與奈米流道間介面之電動捕捉法
2.3.2微孔洞細膜過濾器濃縮法
2.3.3奈米碳管過濾器濃縮法
2.4 分離技術
2.5 偵測單細胞釋放物質之技術
2.5.1 螢光偵測法
2.5.2 質譜儀
2.5.3 表面增強共振拉曼散射法(SERRS)
第三章 實驗設計與製程規劃
3.1 實驗原理
3.1.1 奈米線形成原理
3.1.2 奈米線濃縮機制
3.1.3 電化學偵測原理
3.1.4 場效濃縮機制(DAEKF)
3.2 實驗藥品與配製
3.3 實驗儀器與裝置架設
3.3.1 實驗儀器
3.4 晶片製作流程
3.4.1微晶片流道與偵測電極製作流程
3.4.2 奈米線濃縮區域之製作
第四章 實驗步驟
4.1 時序控制之奈米線濃縮步驟
4.2 奈米線濃縮效果測試
4.3 神經傳導物質之電化學訊號偵測
4.4 PC-12細胞經刺激後所釋出之化學物質之電化學偵測
第五章 結果與討論
5.1. 奈米線濃縮測試結果
5.1.1 奈米線之SEM圖
5.1.2奈米線濃縮效果測試
5.1.3 第三代奈米線濃縮晶片設計與效果測試
5.2 電化學訊號檢測
5.2.1 各濃度神經傳導物質之訊號偵測結果
5.2.2 連續濃度梯度神經傳導物質之偵測結果
5.2.3 場效電場(Field effect)之偵測結果
5.3 類神經細胞PC-12之觀測
第六章 結論
參考文獻

[1] D. D. Carlo and L. P. Lee, "Dynamic Single-Cell Analysis for Quantitative Biology," Analytical Chemistry, vol. 78, 2006,pp. 7918-7925.
[2] D. Bakstad, A. Adamson, D. G. Spiller, and M. R. White, "Quantitative measurement of single cell dynamics," Curr Opin Biotechnol, vol. 23, 2012,pp. 103-9.
[3] T. C. Chao and A. Ros, "Microfluidic single-cell analysis of intracellular compounds," J R Soc Interface, vol. 5 Suppl 2, 2008, pp. S139-50.
[4] Y. Lin, R. Trouillon, G. Safina, and A. G. Ewing, "Chemical analysis of single cells," Anal Chem, vol. 83, 2011, pp. 4369-92.
[5] X. Lu, W.-H. Huang, Z.-L. Wang, and J.-K. Cheng, "Recent developments in single-cell analysis," Analytica Chimica Acta, vol. 510, 2004,pp. 127-138.
[6] M. W. Li and R. S. Martin, "Microchip-based integration of cell immobilization, electrophoresis, post-column derivatization, and fluorescence detection for monitoring the release of dopamine from PC 12 cells," Analyst, vol. 133, 2008, pp. 1358-66.
[7] Wei-Hua Huang, Wei Cheng, Zhen Zhang, "," Analytical Chemistry, vol. 76, 2004,pp. 483-488.
[8] H. Wei, H. Li, S. Mao, and J. M. Lin, "Cell signaling analysis by mass spectrometry under coculture conditions on an integrated microfluidic device," Anal Chem, vol. 83, 2011,pp. 9306-13.
[9] L. Gervais, M. Hitzbleck, and E. Delamarche, "Capillary-driven multiparametric microfluidic chips for one-step immunoassays," Biosens Bioelectron, vol. 27, 2011, pp. 64-70.
[10] Ying-Chih Wang, †, Anna L. Stevens, a. J. Han, and, "million-fold preconcentration of proteins and peptides," Analytical Chemistry, vol. 77, 2005 ,pp. 4293-4299.
[11] J. K. Robert S. Foote, Stephen C. Jacobson, a. J. M. Ramsey, "Preconcentration of Proteins on Microfluidic Devices Using Porous Silica Membranes," Analytical Chemistry, vol. 77, 2005,pp. 57-63.
[12] J.-K. Wu, a, Y.-S. W. , a, C.-S. Y. , b, et al., "Charge-selective gate of arrayed MWCNTs for ultra high-efficient biomolecule enrichment by nano-electrostatic sieving " Biosensors and Bioelectronics vol. 43, 2013,pp. 453-460.
[13] X. Sun, Y. Niu, S. Bi, and S. Zhang, "Determination of ascorbic acid in individual rat hepatocyte cells based on capillary electrophoresis with electrochemiluminescence detection," Electrophoresis, vol. 29, 2008,pp. 2918-24.
[14] M. Z. Bernhard Wolfrum, and Serge Lemay, "Nanofluidic Redox Cycling Amplification for the Selective Detection of Catechol," Analytical Chemistry, vol. 80, 2008.
[15] K. E. Swearingen, W. P. Loomis, B. Kehimkar, B. T. Cookson, and N. J. Dovichi, "Quantification of green fluorescent protein in cellular supernatant by capillary electrophoresis with laser-induced fluorescence detection for measurement of cell death," Talanta, vol. 81, 2010,pp. 948-53.
[16] J. A. D. Lauren M. Ramsay, Oluwatosin Dada, and Norman J. Dovichi, "Femtomolar Concentration Detection Limit and Zeptomole Mass Detection Limit for Protein Separation by Capillary Isoelectric Focusing and Laser-induced Fluorescence Detection," Analytical Chemistry, vol. 81, 2009.
[17] D. M. Omiatek, M. F. Santillo, M. L. Heien, and A. G. Ewing, "Hybrid capillary-microfluidic device for the separation, lysis, and electrochemical detection of vesicles," Anal Chem, vol. 81, 2009,pp. 2294-302.
[18] I. A. Yamboliev, L. M. Smyth, L. Durnin, Y. Dai, and V. N. Mutafova-Yambolieva, "Storage and secretion of beta-NAD, ATP and dopamine in NGF-differentiated rat pheochromocytoma PC12 cells," Eur J Neurosci, vol. 30, 2009,pp. 756-68.
[19] X. T. Zheng, H. B. Yang, and C. M. Li, "Optical detection of single cell lactate release for cancer metabolic analysis," Anal Chem, vol. 82, 2010,pp. 5082-7.
[20] H. Wei, H. Li, D. Gao, and J. M. Lin, "Multi-channel microfluidic devices combined with electrospray ionization quadrupole time-of-flight mass spectrometry applied to the monitoring of glutamate release from neuronal cells," Analyst, vol. 135, 2010,pp. 2043-50.
[21] M. Kaya and M. Volkan, "New approach for the surface enhanced resonance Raman scattering (SERRS) detection of dopamine at picomolar (pM) levels in the presence of ascorbic acid," Anal Chem, vol. 84, 2012,pp. 7729-35.
[22] S. R. Blinks LR, "The Time Course of Photosynthesis as Shown by a Rapid Electrode Method for Oxygen," Proc Natl Acad Sci U S A, 1983,pp. 420-427.
[23] C. Spegel, A. Heiskanen, L. H. D. Skjolding, and J. Emnéus, "Chip Based Electroanalytical Systems for Cell Analysis," Electroanalysis, vol. 20, 2008,pp. 680-702.
[24] D. M. Omiatek, Y. Dong, M. L. Heien, and A. G. Ewing, "Only a Fraction of Quantal Content is Released During Exocytosis as Revealed by Electrochemical Cytometry of Secretory Vesicles," ACS Chem Neurosci, vol. 1, 2010,pp. 234-245.
[25] Y. Zeng, Y. Zhou, L. Kong, T. Zhou, and G. Shi, "A novel composite of SiO2-coated graphene oxide and molecularly imprinted polymers for electrochemical sensing dopamine," Biosens Bioelectron, vol. 45, 2013, pp. 25-33.
[26] C.-Y. Chen, C.-S. Wu, C.-J. Chou, and T.-J. Yen, "Morphological Control of Single-Crystalline Silicon Nanowire Arrays near Room Temperature," Advanced Materials, vol. 20, 2008,pp. 3811-3815.
[27] J. B. Z. a. R. M. W. Kirk T. Kawagoe, "Principles of voltammetry and microelectrode surface states," Journal of Neuroscience Methods, vol. 48, 1993,pp. 225-240.
[28] R. G. Wu, C. S. Yang, C. K. Lian, C. C. Cheing, and F. G. Tseng, "Dual-asymmetry electrokinetic flow focusing for pre-concentration and analysis of catecholamines in CE electrochemical nanochannels," Electrophoresis, vol. 30, 2009,pp. 2523-31.