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研究生:洪羚軒
研究生(外文):Ling-Hsuan Hung
論文名稱:利用微環境影響黃色嵌紋病毒於感測表面之著附效果
論文名稱(外文):Increasing adhesive density of TYMV on sensors'' surface by controlling microfluidic enviornment
指導教授:吳嘉哲
指導教授(外文):Chia-Che Wu
學位類別:碩士
校院名稱:國立中興大學
系所名稱:機械工程學系所
學門:工程學門
學類:機械工程學類
論文種類:學術論文
論文出版年:2010
畢業學年度:98
語文別:中文
論文頁數:90
中文關鍵詞:混沌形式微環境渦度浸泡法MUANCD-4TYMV
外文關鍵詞:chaotic flowmicroenvironmentCOMSOL MultiphysicMUANCD-4TYMVdipping method
相關次數:
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目前生物檢測方式中,利用抗體與抗原之間的辨識與連結之方法已普遍被用於各方面之檢測,例如將抗體以生物技術之方法固定於感測表面,然後利用抗體與抗原之間的辨識性與專一性,讓抗體經由辨識後,能自動抓取待測液體中的抗原(例如病毒、細菌或是過敏原等)。當抗原一旦與感測區上之抗體連結之後,藉由量測感測區域於感測前和感測後之物理或化學性質的改變,來判定待測液中是否含有特定抗原。常見的方法,可利用光學、電性等方法,對感測區域進行分析量測。
然而,感測區域上的辨識元件(抗體),其覆蓋面積與覆蓋均勻度,對於感測器之準確性(Accuracy)與靈敏度(Sensitivity)有很大的影響。辨識層於感測區之覆蓋比率越高、均勻性越好,則感測區域能辨識與抓取抗原的能力也就越好。而當得到品質優良的辨識層後,另一個問題是,如何讓待測液中的抗原,充分著附於感測區域上。當感測區上之抗原數量極少或是分佈不均時,常常導致量測不到訊號的變化或是訊號與雜訊過於接近而無法判讀分析結果,為了避免以上的情況,則需使抗原著附於感測區域之數量與均勻性越高越好,以利量測與分析感測區域在感測前後之物理、化學變化量,並且可降低感測器可感測之最低濃度與待測樣本所需之體積。
本研究為了改善上述之問題,利用微環境中的結構使微流體產生多方向之旋轉流動,讓感測區域的待測液體形成混亂的流動,藉此增加辨識層以及抗原著附於感測區域之覆蓋率與均勻性。首先在微環境之設計上,藉由有限元素模擬軟體(COMSOL Multiphysics)之輔助,進行微環境之流場、流線與渦度之模擬,由模擬結果進行參數修正,以得到能產生預期目標之微環境形式,並且利用螢光流線實驗來驗證模擬結果。接著將以NCD-4螢光分子為檢體,對MUA自我組裝分子膜進行覆蓋率與均勻性之檢測,並與傳統浸泡方式進行比較,實驗結果顯示,傳統浸泡法之覆蓋率為33.37%;混沌流形式的MUA分子層,其覆蓋率可達96.3%,增加幅度約為188%。最後,以TYMV植物病毒為抗原,實際進行抗原著附於感測區之實驗,並與傳統浸泡方式比較,實驗結果顯示,傳統浸泡法之覆蓋率則為4.23%;混沌流形式之微環境,其TYMV於感測區之覆蓋率為70.1%,增加幅度約為1557%。其中,以浸泡法所得之TYMV單位時間內之覆蓋率為7.05×10-3 %/min;混沌流形式之微環境則為50.7 %/min,為傳統浸泡法之7102倍,此結果顯示,混沌流形式之微環境不僅提升MUA分子層與TYMV抗原之著附覆蓋率,並且大幅縮短TYMV著附至感測區域所需之間,對於未來進行快速生物檢測將有很大的幫助。


Currently, antibody-antigen reaction have been used in biological detection. Antibody can identify specific antigen from samples. Similarly, only specific key can open the specific lock. One of biological detection methods, antibodies are first affixed to the sensing surface in microenvironment. When samples are designed to pass the sensing surface, antibodies will capture the antigens in samples.
Researchers typically uses either optical or electrical measurement to examine the sensing surface for specific antigen. In optical measurement, antigens were labeled fluorescent dyes which can be used to estimate the amount of antigen-antibody on the sensing surface. In electrical measurement, the impedance measurement of the sensing surface can also be used to determine the quantity of antibody-antigen. For optical or electrical measurement, it is very important to have dense antigen-antibody on the sensing surface. Poor density of antibody-antigen will results to poor sensitivity of biological detection.
The purpose of this research is to enhance the density of antibody-antigen on sensing surface by controlling microenvironment. Chaotic flow of samples was produced by the structure and the conditions of microenvironment. Finite element analysis – COMSOL Multiphysic, was used to compute the velocity field, curl field and streamline of samples in microenvironment. Fluorescent particles was used to show streamline of samples experimentally. Experimental result was compared to simulation one. Finally, Turnip Yellow Mosaic Virus was used to be the specific antigens experimentally. Experimental results show that the density of TYMV detected by chaotic flow was 16.5 times larger than the density detected by dipping method. The duration of experiment by chaotic flow was 0.0023 times less than the duration by dipping method. This research provided a simple and efficient design that benefit rapid and real-time detection.


致謝 I
摘要 II
Abstract IV
目錄 V
圖目錄 VII
表目錄 X
第一章 緒論 1
1.1 研究動機 1
1.2 研究目標 9
1.3文獻回顧 10
1.4 論文架構 13
第二章 TYMV於微環境之相關理論 14
2.1 分子生物學 (Molecular Biology) 14
2.1.1 病毒結構 15
2.1.2 抗原–抗體反應(Antigen–Antibody interaction) 20
2.2 分子動力學 (Molecular dynamics) 23
2.2.1 擴散效應與布朗運動 23
2.2.2 聚集現象 29
2.2.3 靜電作用(Electrostatic interaction) 30
2.3 感測原理 (Sening theory) 32
2.3.1 TYMV著附原理 32
2.3.2 共軛焦顯微鏡(Confocal microscope) 33
2.4 流體力學 (Hydrodynamics) 35
2.4.1 Reynolds number與Peclect number 35
2.4.2 流線(Steamline)與渦度(Curl) 38
第三章 微環境形式設計與分析 40
3.1 不同形式的微環境 40
3.1.1 病毒靠擴散的作用方式 41
3.1.2 病毒靠流力傳輸的作用方式 43
3.2 流場、流線與渦度之模擬結果 45
3.3 螢光流線實驗 61
3.3.1 微環境製程 61
3.3.2 微環境螢光流線 62
第四章 辨認層介紹與製備 66
4.1 MUA自我組裝分子層 66
4.2 EDC / NHS配體層 67
4.3 NCD-4螢光檢體介紹 69
4.4 Alexa Fluor 594螢光介紹 72
第五章 著附結果與討論 74
5.1 檢驗MUA自我組裝分子層 74
5.1.1 實驗控制組 74
5.1.2 傳統浸泡方式 75
5.1.3 層流形式微環境 76
5.1.4 混沌流形式微環境 78
5.2 TYMV著附實驗之定量分析 80
5.2.1傳統浸泡方式 80
5.2.2層流形式微環境 80
5.2.3混沌形式微環境 81
5.2.4 TYMV覆蓋率與著附效率比較 82
第六章 結論與未來展望 85
6.1 結論 85
6.2 未來展望 86
參考文獻 87



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