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研究生: 林湘庭
Xiang-Ting Lin
論文名稱: 開發感溫型透明質酸/明膠微球應用於幹細胞擴增培養系統
Development of thermosensitive hyaluronic acid/gelatin microspheres for stem cell expansion culture system
指導教授: 蔡協致
Hsieh-Chih Tsai
口試委員: 林宣因
Suian-Yin Lin
陳玉暄
Yu-Shuan Chen
高震宇
Chen-Yu Kao
學位類別: 碩士
Master
系所名稱: 應用科技學院 - 應用科技研究所
Graduate Institute of Applied Science and Technology
論文出版年: 2022
畢業學年度: 111
語文別: 中文
論文頁數: 91
中文關鍵詞: 微載體培養系統溫度敏感型共聚物透明質酸明膠細胞貼附率細胞脫附
外文關鍵詞: Microcarrier culture systems, thermosensitive copolymer, hyaluronic acid, gelatin, cell adhesion rate, cell desorption
相關次數: 點閱:267下載:0
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致謝 IV 摘要 V ABSTRACT VI 目錄 VIII 圖目錄 XII 表目錄 XV 第一章 前言 1 第二章 文獻回顧 2 2.1 微載體 2 2.1.1 微載體的定義 2 2.1.2 微載體的類型 3 2.1.3 微載體的表面性質 5 2.1.4 溫敏性微載體 7 2.2 微載體的製備方法 8 2.2.1 乳化反應 9 2.2.2 親水親油平衡(HLB) 10 2.2.3 表面活性劑 11 2.3 微載體的應用 12 2.3.1 幹細胞治療 12 2.3.2 組織工程 13 2.3.3 疫苗生產 14 2.4 細胞培養 15 2.4.1 二維細胞培養 15 2.4.2 三維細胞培養 15 2.5 細胞的脫貼附機制 17 2.5.1 細胞貼附機制 17 2.5.2 酶解法分離細胞 17 2.5.3 非酶解法分離細胞 17 2.6 刺激響應性聚合物 19 2.6.1 熱響應聚合物 21 2.6.2 溫敏性高分子PNIPAM 22 2.6.3 親水性單體Acrylic acid(AAc) 22 2.6.4 明膠 23 2.6.5 EDC交聯劑 24 2.7 研究動機 25 第三章 實驗方法 26 3.1 實驗流程 26 3.2 實驗藥品 27 3.3 實驗設備 29 3.4 分析儀器 30 3.5 實驗方法 31 3.5.1 溫度敏感型共聚物P(NIPAM-AAc)之合成 31 3.5.2 GLA微球之合成 32 3.5.3 G-HA微球之合成 32 3.5.4 G-HA-L微球之合成 33 3.6 溫度敏感型共聚物之鑑定與分析 34 3.6.1 核磁共振光譜(1H-NMR)分析 34 3.6.2 分子量測量分析 34 3.7 微球物化性質分析 34 3.7.1 粒徑分佈測量 34 3.7.2 穩定度測試 34 3.7.3 膨潤度及含水率測量 34 3.7.4 溶解行為測試 35 3.8 細胞實驗 36 3.8.1微球之細胞存活率 36 3.8.2 微球之細胞貼附率 36 3.8.3 微球之細胞脫附測試 37 3.8.4 微球之細胞貼附情形與活性試驗 37 第四章 結果與討論 38 4.1 溫度敏感型共聚物合成 38 4.1.1 核磁共振光譜(1H-NMR)鑑定與分析 38 4.1.2 凝膠滲透色譜(GPC)分子量測量 40 4.1.3 最低臨界溶解溫度(LCST)量測 41 4.2 明膠微球分析與鑑定 42 4.2.1 衰減全反射傅里葉變換紅外光譜儀(ATR-FTIR)分析 42 4.2.2 拉曼光譜儀(Raman Spectroscopy)分析 45 4.2.3 粒徑分佈測量 49 4.2.4 穩定度測試 50 4.2.5 掃描式電子顯微鏡分析(SEM) 51 4.2.6 膨潤度及含水率測量 52 4.2.7 溶解行為測試 53 4.2.8 流變儀分析 54 4.3 生物相容性試驗 55 4.3.1 溫度敏感型聚合物P(NIPAM-AAc)之細胞存活率 55 4.3.2 微球之細胞存活率 56 4.4 細胞脫貼附試驗 57 4.4.1 微球之細胞貼附情形與活性試驗 57 4.4.2 微球之細胞貼附率 58 4.4.3 Dispase® II之細胞脫附測試 61 4.4.4 LCST之細胞脫附測試 62 第五章 結論 67 第六章 參考文獻 69

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