臺灣博碩士論文加值系統

本研究利用微機電系統製程技術之灌注成形法以及微壓印技術，完成聚二甲基矽氧烷(Polydimethylsiloxane, PDMS)濃度晶片之製備，以應用於老鼠肌母細胞(C2C12)對血小板衍生生長因子(Platelet-derived growth factors, PDGF)最適濃度之研究。並成功的製備一個微濃度晶片，可同時間檢測六種蛋白質濃度對細胞遷移之效果，蛋白質檢測實驗方法首先是細胞外基質(Extracellular matrix, ECM)最適濃度選取以利後續實驗進行，探討不同體積百分濃度之ECM(100%、80%、50%、20%、10%、5%、2%、0%)選出最適合老鼠肌母細胞生長之濃度，加速細胞生長速度以利實驗觀察，然後在晶片中固定細胞培養區域並利用微壓印技術壓印出不同濃度之PDGF，觀察其細胞遷移效應之影響；另外經實驗驗證在10%的濃度下細胞生長快速且成本最低，而肌母細胞對於PDGF之最適濃度為10~100 μg/mL，當PDGF濃度過高或過低於此最適濃度時生長效果皆沒有比此範圍濃度好。故本研究所建立的濃度晶片系統，不但可解決傳統藥物測試上無法觀測細胞遷移效果且不能於同一實驗區內進行多種濃度測試的問題，在未來將有助於藥物濃度測試及藥物過敏測試等應用領域的發展。

This thesis presents a new microfluidic Polydimethylsiloxane (PDMS) chip with pre-concentration unit for determining the optimal Platelet-derived growth factors (PDGF) concentration in mouse myoblast cells. First, we determined the optimal concentration of Extracellular matrix (ECM) to accelerate the growth of mouse myoblast cells. Then, we fixed the cells in the cultured area and used the micro-imprinting chip to emboss different concentrations of PDGF. When the concentration of PDGF was between 10 μg/mL and 100 μg/mL, the growth effect was optimal. Therefore, the proposed chip not only solves the efficiency problem of conventional drug test devices, but also can text various drug concentrations in the same region on one chip.

摘要 I
EXTENDED ABSTRACT III
誌謝 IX
目錄 X
圖目錄 XIII
第一章 緒論 1
1-1 文獻回顧 3
1-1-1 微圖案於生物培養之技術 5
1-1-2 細胞遷移 11
1-1-2-1 細胞遷移的重要分子 13
1-2 血小板衍生生長因子簡介 15
1-3 細胞外基質簡介 16
1-4 研究動機與目的 17
1-5 研究架構 19
第二章 晶片之設計與製作 20
2-1 光罩設計與製作 20
2-1-1 濃度測試晶片光罩設計與製作 20
2-1-2 微壓印頭光罩設計與製作 22
2-2 THB-151N製程技術與微結構製作 24
2-2-1 THB-151N製程技術 24
2-2-2 微結構製作 25
2-3 PDMS灌注成形技術及翻製流程 32
2-4 PDMS晶片裝置製作 35
第三章 實驗與研究方法 36
3-1 實驗藥品及其配製 36
3-1-1 實驗藥品 36
3-1-2 藥品配製 37
3-2 細胞培養與收集計數 38
3-2-1 細胞繼代培養 40
3-2-1-1 細胞解凍 41
3-2-1-2 高速離心機 42
3-2-2 細胞計數 43
3-3 實驗儀器 46
3-3-1 倒立式螢光光學顯微鏡 46
3-3-2 表面粗度儀 47
3-3-3 真空抽氣系統 48
3-3-4 倒立式螢光光學顯微鏡(動態記錄) 49
3-4 實驗方法 50
3-4-1 細胞於ECM最適濃度之實驗 51
3-4-2 PDGF濃度對細胞遷移之趨向性實驗 52
第四章 結果與討論 54
4-1 壓印ECM間距實驗結果 54
4-2 ECM最適濃度實驗結果 56
4-3 PDGF與細胞間距實驗結果 57
4-4 PDGF濃度對細胞遷移之趨向性實驗 61
4-4-1 PDGF濃度(0.01~1000 μg/mL) 61
4-4-2 PDGF濃度(5~1000 μg/mL) 64
4-4-3 PDGF濃度(1~50 μg/mL) 66
4-5 PDGF濃度對細胞遷移之趨向性實驗(動態記錄) 69
第五章 結論與建議 71
5-1 結論 71
5-2 建議 72
參考文獻 74

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