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研究生:黃一倉
論文名稱:細胞捕捉微流體元件整合雷射加熱在藥物作用後細胞DNA局部熱變性之研究
論文名稱(外文):Cell Capture Microfluidic Device Integrated with Laser-induced Heating for Local Denaturation of DNA Fibers from a Cell Treated with Drugs
指導教授:洪敏勝
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:100
語文別:中文
中文關鍵詞:單一細胞捕捉DNA拉伸DNA熱變性
外文關鍵詞:single cell captureDNA extendingDNA denaturation
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本研究結合微流體元件與雷射加熱系統,開發單一細胞捕捉與藥物作用後細胞DNA局部熱變性之微流體系統。本研究使用之微流體元件以PDMS製作完成,流道之設計主要包含細胞捕捉、釋放區及DNA拉伸與分析區。細胞捕捉及釋放之原理為以主、支流道流體體積流率差異捕捉單一細胞,利用雷射聚焦於雷射加熱區,使水溶液因高溫產生氣泡而釋放細胞。DNA拉伸與分析區則用於裂解被釋放的細胞,並進行細胞DNA拉伸與DNA局部位置之熱變性。研究結果顯示,主流道流速為1.2×10^4μm/s時細胞因受到較大的流體壓力作用,細胞產生變形而通過微結構,降低細胞捕捉效率。主流道流速為80μm/s時細胞受到較小的流體作用力,被微結構捕捉、固定的細胞不會變形,捕捉效率因而增高。當雷射功率為164mW、主流道流速≦ 80μm/s時,雷射加熱產生的氣泡可將細胞推離被捕捉的位置而釋放細胞。當細胞被釋放後,沿著主流道往DNA分析區流動並被固定,注入1% SDS溶液後可裂解細胞,同時使細胞DNA釋放於溶液中而被拉伸,並固定於下游所設計之圓柱結構。最後利用雷射加熱固定於圓柱結構間的DNA,當雷射功率為47mW且照射30秒時,可達到DNA局部熱變性之目的。
This study combines a microfluidic device and laser-induced heating system to develop a single cell capture that has been treated with drugs and partial cellular DNA denaturation microfluidic system. The microfluidic device utilized in this study consists of PDMS. The flow channel design consists mainly of cell capture and release, and DNA stretching and analysis zone. The concept of single cell capture is use the flow rate different between main flow channel and branch channel. The laser is focused on the laser heating zone to generate a bubble to release the captured cell. Finally, DNA stretching and partial cellular DNA denaturation is demonstrated in DNA stretching and analysis zone after cell lysis. The results show that larger hydrodynamic stresses act on the cell and move cell to flow through the micro-structure at high flow rate of 1.2×10^4μm/s, reducing the efficiency of cell capture. At low flow rate of 80μm/s, slight hydrodynamic effect on the cell and the cell can be captured; the capture efficiency is thus increased. In addition, when velocity in main flow channel is less than 80μm/s and laser power is 164mW, the cell can be released by expanded bubble after laser irradiation. The released cell flows into DNA analysis zone and is captured by micro-structure. The cell lysis and DNA extending are achieved after 1% SDS injection. Finally, the laser is used to irradiate the DNA fibers anchored between micro-pillars. When the laser power is 47mW, 30 seconds of irradiation, the partial DNA denaturation can be observed.
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細胞捕捉微流體晶片 2
1.1.2細胞分析晶片 5
1.2 研究目的 10
第二章 理論基礎 11
2.1 細胞捕捉理論 11
2.2 雷射加熱理論 14
第三章 實驗設備與方法 16
3.1 實驗設備與系統 16
3.2 實驗樣本與藥品 18
3.3微流體元件製作 19
3.4 實驗方法 24
第四章 結果與討論 27
4.1細胞捕捉 27
4.2以雷射加熱釋放捕捉的細胞 31
4.2.1雷射聚焦位置 33
4.2.2雷射功率對氣泡產生的影響 33
4.2.3不同流速對於氣泡產生的影響 37
4.2.4以雷射加熱釋放細胞 40
4.3藥物作用後細胞DNA之拉伸 46
4.3.1秋水仙素對細胞的影響 47
4.3.2細胞固定與DNA拉伸 48
4.3.3 DNA之局部熱變性 50
第五章 結論與建議 52
5.1結論 52
5.2 建議 53
參考文獻 54
附錄 雷射通過不同物鏡後之功率 58

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