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研究生: 廖士慶
Shih-Ching Liao
論文名稱: 設計與製作回授控制壓力式生物反應器以探討壓力對骨髓幹細胞增生與型態之影響
Design of an Automatic Control Hydrostatic Pressure Bioreactor for the Investigation of Bone Marrow Stem Cell Growth
指導教授: 黃幸宜
"none"
鍾志昂
Chih-Ang Chung
口試委員:
學位類別: 碩士
Master
系所名稱: 工學院 - 機械工程學系
Department of Mechanical Engineering
畢業學年度: 97
語文別: 中文
論文頁數: 65
中文關鍵詞: 組織工程生物反應器
外文關鍵詞: Tissue enginnering, Bioreacotr
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  • 本研究主旨為設計一套壓力式生物反應器,為了得到穩定且正確的壓力值,我們架構了回授控制系統,使得生物反應器可施加定壓或循環壓力。測試結果顯示,壓力值最大可達到5 MPa,頻率可達1 Hz。並利用無菌採樣袋成功節省實驗中培養液使用量,經由實驗證明無菌採樣袋內環境適合細胞培養。
    我們使用此生物反應器給予大鼠骨髓幹細胞 (RM1) 定常壓力 (0.4、0.55、0.7、1 MPa),其中1 Ma實驗,施以二組不同的加壓斜率到達定值1 MPa,測試加壓斜率對大鼠髓幹細胞生長速率的影響。實驗結果證明初始加壓斜率過大會抑制大鼠骨髓幹細胞生長。往後可利用此回授控制壓力式生物反應器給予不同壓力式形刺激細胞,觀察壓力對細胞基因表現的影響。


    The study design a pressure bioreactor, in order to steady and correct pressure value, we frame automatic control hydrostatic system that can provide constant and time-periodic pressure for in vitro cell cultures. The systems’ performance was stable at a constant hydrostatic pressure up to 5 MPa and cyclic hydrosatatic pressure up to 1 Hz. Cells were putting in the whirl-Pak Sample Bags, which could largely reduce the amount of culture media and prevent contamination.
    We tested the rat bone marrow stem cells (RM1) by applying 0.4, 0.55, 0.7 and 1 MPa of static pressure at the bioreactor. We study how the pressure slope in time may influence the cell growth by, we changed the pressure slope for the 1 MPa case. The results show it is likely to be the pressure slope rather than the final static pressure value that inhibited the RM1 growth. The cells growth apparently slowed down when the pressure slopes were large than 700 KPa/s. The bioreactor was proved to be available in cells culture and it will be applied to test cells for gene expressions in the future.

    目錄 中文摘要……………………………………………………………………………. i 英文摘要…………………………………………………………………………….ii 目錄………………………………………………………………………………....iii 圖目錄……………………………………………………………………...………. vi 第一章 緒論 1.1 前言………………………………………………………………………...1 1.2 研究動機…………………………………………………………………...2 1.3 文獻回顧…………………………………………………………………...3 1.3.1 壓力對細胞培養的影響……………………………………………..3 1.3.2 壓力式生物反應器的設計…………………………………………..5 1.3 實驗構想…………………………………………………………………...6 第二章 生物反應器之設計與製作 2.1 設計前言…………………………………………………………………...7 2.2 細胞培養部分……………………………………………………………...8 2.2.1 培養室設計…………………………………………………………9 2.2.2 無菌採樣袋…………………………………………………………9 2.3 壓力驅動部分…………………………………………………………… 10 2.3.1 自動控制回授系統………………………………………………..12 2.3.1.1 回授控制硬體架構……..……………………………….....12 2.3.1.2 回授控制軟體……………………………………………...13 2.4 壓力驅動性能測試……………………………………………………….16 第三章 實驗方法 3.1 細胞載體………………………………………………………………….20 3.2 細胞來源………………………………………………………………….21 3.3 玻片消毒與細胞種植…………………………………………………….21 3.4 熱封無菌採樣袋………………………………………………………….22 3.5 無菌採樣袋細胞相容性測試…………………………………………….23 3.5.1 無菌採樣袋生長環境測試………………………………………..23 3.5.2 更換無菌採樣袋測試……………………………………………..27 3.6 壓力式生物反應器實驗步驟…………………………………………….30 3.6.1 生物反應器組裝…………………………………………………..30 3.6.2 生物反應器實驗流程……………………………………………..31 3.7 實驗結果分析方法……………………………………………………….32 3.7.1 顯微鏡觀察和拍攝………………………………………………..32 3.7.2 細胞計數…………………………………………………………..33 第四章 實驗結果 4.1 實驗一 (0 KPa)…………………………………………………………..34 4.2 實驗二 (1 MPa)…..………………………………………………………37 4.3 實驗三 (0.4 MPa).………………………………………………………..41 4.4 實驗四 (0.7 MPa)….……………………………………………………..45 4.5 實驗五 (0.55 MPa)…...…………………………………………………..49 4.6 實驗六 (1 MPa)…..………………………………………………………53 第五章 結論與未來展望…………………………………………………………57 參考文獻……………………………………………………………………………..59 附錄 實驗試藥及儀器………………………………………………………………..64

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