本研究開發一新的檢測系統，使用微奈米濃縮晶片對生物樣本進行預濃縮後，利用氣閥捕捉濃縮區塊，再進行布朗運動的檢測分析。主要的微奈米流體濃縮晶片，是使用Nafion奈米薄膜作為離子選擇性通道，又可稱為濃縮器(precon- centrator)，施加一電位差於具有離子選擇性(ion-selective)效果的Nafion薄膜上可產生濃度極化效應(ion concentration polarization)，於高電位處產生離子空乏區(ion depletion region)。此時施加偏壓可產生第二種電滲透流(electroosmosis of second kind)，可在高電位處的空乏區旁產生原本樣本濃度的105~106倍的濃縮區塊(preconcentration plug)，可以大幅降低濃度的生物樣本的偵測下限。達到低濃度生物標記(biomarker)的偵測，使用氣閥(pneumatic valve)將濃縮區塊捕捉後，結合布朗運動，利用抗原與抗體間之專一性(Specific Relationship)辨認作為檢測機制。
　　本文詳述使用Nafion奈米薄膜濃縮晶片的設計與驗證，包括Nafion的導電率與晶片電阻模型的驗證、以及氣閥捕捉濃縮區塊的成效。使用LabVIEW架設迴路電流監測系統觀測電流與濃縮之間的特性，並以微粒子追蹤測速儀(micro-Particle Pracking Velocimetry, micro-PTV)進行布朗運動的量測與分析。
　　本研究所開發的檢測方法具有降低生物樣本偵測下限，提高檢測靈敏度的能力，因此將來可以使用在不同的低濃度生物樣本上，以利後端的生物檢測。

In this thesis, we have developed a novel detection method based on capturing preconcentrated plug preconcentrated by preconcentration chip and Brownian motion detection. Applying voltage drop to the preconcentrator which use Nafion as ion-selective membrane can generate ion concentration polarization (ICP) effect. The electroosmosis of second kind which generate by applying the bias voltage, force bulk solution to aggregate the preconcentration plug near the depletion region. Using the Brownian motion which analyze the characteristic of antigen-antibody interactions as detection mechanism to detect the preconcentration plug captured by pneumatic valve.
　　The design and the validation methods and processes of nanofluidic preconcentration chips and pneumatic valve were proposed starting from validation of resistive models, test of ion-selective membrane and test of Brownian motion by micro-Particle Tracking Velocimetry (micro-PTV), to get the onset of detection mechanism.
　　In summary, the presented nanofluidic preconcentrator and pneumatic valve demonstrates biological sample can be preconcentrated in short time and the plug can be captured by valve. With the analysis of loop currents and Brownian motion detection, various low biological sample could be demonstrated in the future.

致謝 i
摘要 ii
Abstract iii
目錄 iv
表目錄 vii
圖目錄 viii
第一章 導論 1
1-1前言 1
1-2 研究動機與目的 1
1-3 研究方法 2
1-4 論文架構 2
第二章 文獻回顧 3
2-1生物晶片感測文獻回顧 3
2-1-1微全程分析系統（Micro Total Analysis Systems, μTAS） 3
2-1-2免疫分析法(Immunoassay) 3
2-1-3樣本前處理之欲濃縮技術 4
2-2微奈米流體濃縮晶片文獻回顧 5
2-2-1 微奈米流體晶片之發展 5
2-2-2奈米流體晶片製程 10
2-3布朗運動文獻回顧 13
2-4微粒子影像測速儀/粒子追蹤測速儀文獻回顧 14
第三章 實驗理論及技術背景 17
3-1微奈米濃縮晶片之濃縮原理 17
3-1-1電雙層 17
3-1-2離子的區域性空乏與濃縮現象 20
3-1-3離子選擇性薄膜之電壓與電流的S曲線 24
3-1-4預濃縮機制 (Mechanism of preconcentration) 25
3-2以 Nafion離子選擇性材料為奈米流道 26
3-3微奈米濃縮晶片之免疫分析原理 27
3-3-1生物分子相互作用分析法 27
3-4 布朗運動數學模型 28
3-4-1 愛因斯坦關係式 29
3-4-2 朗之文方程式 31
第四章 實驗設備與實驗方法 33
4-1 濃縮晶片的設計與製程 34
4-1-1製程方法 34
4-2微奈米流體濃縮晶片之量測系統 41
4-2-1 倒立式螢光顯微鏡量測系統 41
4-2-2 LabView架設迴路電流量測系統 42
4-3 NafionR之導電率量測與驗證 44
4-4 微粒子影像與微粒子追蹤測速儀原理 46
4-4-1 微粒子影像/追蹤測速實驗設備 48
4-5 奈米粒子表面之抗體修飾 52
4-5-1螢光粒子之選用 52
4-5-2奈米粒子表面之抗體修飾流程 52
第五章 實驗結果與討論 55
5-1微奈米流體濃縮晶片結果與討論 55
5-1-1利用觀測迴路電流驗證濃縮晶片機制 55
5-1-2濃縮機制現象的觀察與討論 58
5-1-3蛋白質濃縮倍率之亮度分析 59
5-2氣閥捕捉特定區塊之分別驗證 62
5-2-1捕捉區塊後布朗運動檢測 63
5-2-2捕捉區塊後的濃縮效果 64
第六章 總結與未來展望 67
6-1 總結 67
6-2 未來展望 68

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