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研究生:邵毅東
研究生(外文):Yi-Dong Shao
論文名稱:快速原生質體分離與回收微流體平台之開發
論文名稱(外文):Microfluidic Platform for Rapid Separation and Recoveryof Protoplasts
指導教授:洪敏勝
指導教授(外文):Min-Sheng Hung
學位類別:碩士
校院名稱:國立嘉義大學
系所名稱:生物機電工程學系研究所
學門:工程學門
學類:機械工程學類
論文種類:學術論文
畢業學年度:102
語文別:中文
中文關鍵詞:蝴蝶蘭原生質體分離微流體元件螺旋流道
外文關鍵詞:PhalaenopsisProtoplasts separationMicrofluidic deviceSpiral channel
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本研究使用微製程技術,利用微結構製程與少量藥劑開發即時蝴蝶蘭(Phalaenopsiahs)原生質體(protoplast)分離與回收微流體系統。為了達到分析生物樣本的目的,使用一般的生化方法(例如沉澱、離心、與試劑進行反應及分離操作)常因為生物樣本在試管中遺失,而無法獲得大量且完整的樣本,使得後續研究不容易進行。本研究設計並製作螺旋流道回收不同尺寸的微粒與細胞,並用於分離、回收蝴蝶蘭葉片之原生質體。實驗中同時探討不同流量對於微粒與細胞回收之影響,最後將微流體元件用於分離、回收原生質體。實驗結果顯示,由於直徑6μm微粒的Lift force小於 Dean force,粒子於外側流道流動,外側出口有較高的回收率。直徑10μm微粒的Lift force約等於Dean force,微粒於流道中間流動,內、外側出口的回收率約略相等。直徑15μm的U937細胞Lift force大於 Dean force,細胞偏向內側流動,內側出口有較高的回收率。而當用於原生質體分離與回收時,由於溶液中含有細胞壁等雜質,原生質體分離之效果不佳。
致謝......................................................................................................................i
摘要.....................................................................................................................ii
Abstract..............................................................................................................iii
目錄....................................................................................................................iv
圖目錄................................................................................................................vi
表目錄..............................................................................................................viii
符號說明............................................................................................................Ix
第一章 前言.....................................................................................................1
1.1研究背景.............................................................................................1
1.1.1細胞純化與回收.........................................................................2
1.2研究目的.............................................................................................5
第二章 基礎理論...........................................................................................6
2.1植物細胞..........................................................................................6
2.2曲面流體動力分析.............................................................................7
第三章 實驗材料與方法.................................................................................10
3.1實驗設備與材料................................................................................10
3.1.1 實驗系統..................................................................................10
3.1.2 實驗樣本與藥品......................................................................11
3.2 微流體元件製作.............................................................................13
3.2.1負光阻製程................................................................................13
3.2.2 PDMS製程................................................................................14
3.2.3 氧電漿貼合..............................................................................15
3.3 實驗方法.........................................................................................16
第四章 結果與討論.........................................................................................20
4.1 PDMS微流體元件...........................................................................20
4.2流場模擬...........................................................................................22
4.2.1 數值模擬運算..........................................................................22
4.2.2 數學模型..................................................................................22
4.2.3流場模擬計算............................................................................23
4.3 微粒與細胞分離.............................................................................30
4.3.1 直徑6μm粒子..........................................................................30
4.3.2 直徑10μm粒子........................................................................32
4.3.3 直徑15μm U937細胞..............................................................34
4.4 以微流體元件回收原生質體.........................................................37
第五章 結論與建議.........................................................................................41
5.1 結論.................................................................................................41
5.2 建議.................................................................................................42
參考文獻...........................................................................................................43
附錄一酵素液調配...........................................................................................47
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