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研究生:簡弘量
研究生(外文):Hung-Liang Chien
論文名稱:以奈米模板製備導電高分子奈米探針並應用於免疫分析
論文名稱(外文):Fabrication of polyaniline nanowires for immunosensing using porous anodic aluminum oxide template
指導教授:盧燈茂盧燈茂引用關係莊承鑫莊承鑫引用關係
指導教授(外文):Deng-Maw LuCheng-Hsin Chuang
學位類別:碩士
校院名稱:南台科技大學
系所名稱:機械工程系
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:102
畢業學年度:101
語文別:中文
論文頁數:79
中文關鍵詞:陽極氧化鋁聚苯胺免疫分析
外文關鍵詞:aluminum oxide (AAO)PANIImmunosensing
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本論文係利用多孔性陽極氧化鋁作為奈米模仁,以電化學方式製備導電高分子聚苯胺奈米線,作為免疫分析的晶片,透過電化學阻抗頻譜來分析膀胱癌T24之裂解中,與Galectin-1抗體所相對應之特性蛋白質的存在。本研究再以陽極氧化製程,製作出孔徑約130奈米及約138微米厚的多孔性氧化鋁模仁。接著為氧化鋁模板濺鍍一層導電白金層作為電化學沉積所需要的導電層。完成白金濺鍍的模板使用三極式電化學工作平台進行恆電位法沉積聚苯胺奈米線,再以氫氧化納蝕刻陽極氧化鋁模仁表面,使一片分散性良好的聚苯胺奈米探針顯露出來,奈米探針在不團簇的情況下皆能夠獨自提供感測面域,因此可獲得超高的表面積,致提升其感測之靈敏性。為了測試聚苯胺奈米探針具有檢測的能力,所以在本實驗中先使用0.1μM的牛血清白蛋白(BSA-AF488)與其專一性鍵結的抗體(Antibody-BSA)進行免疫分析,透過循環伏安法(Voltammetry,CV)以及螢光反應(Fluorescent Detected)初步的驗證其感測能力。
根據實驗結果的觀察,在螢光檢測上可明顯的分辨出有無BSA蛋白質鍵結之螢光差異;而從循環伏安中的電位-電流曲線圖中也清楚的顯示出其曲線的變化。從螢光的差異性及CV曲線圖的變化等結果,可顯示出聚苯胺奈米探針之免疫分析能力。接著再更進一步的利用電化學阻抗頻譜(Electrochemical Impedance Spectroscopy, EIS)來分析膀胱癌T24裂解液中之特性蛋白質的存在,並比較在不同酸鹼值之環境下的阻抗的變化。根據最後的結果顯示,聚苯胺奈米探針的免疫晶片成功的達到免疫分析的檢測。
In this study, we have developed an immunosensor with well-separated PANI nanofibers by using anodic aluminum oxide (AAO) template for enlarging the surface-volume ratio and enhancing the sensitivity of impedance sensing. The AAO template with pore diameter of 120 nm and thickness of 138 um was made by anodization process and subsequently evaporated with platinum metal layer on the backside for further electrochemical deposition of PANI nanofibers. The PANI nanofibers were synthesized with 1M hydrochloric acid with 0.3M aniline by potential-static electrochemical deposition. The well-separated PANI nanofibers were finally released by 1M NaOH etching for a few minutes. As the results, the well-separated PANI nanofibers were expected to provide more surfaces for immunoreaction and better electron transfer for impedance sensing. For the demonstration of immunosensing, 0.1 μM labeled BSA-AF488 protein was successfully detected based on fluorescence measurement after the PANI nanofibers were conjugated with BSA antibodies. Furthermore, the immunosensor was employed to detect the protein expression of bladder cancer cell lysate (grade III:T24) by antibody Galectin-1 in different pH value buffer for determination of isoelectric point (pI). According to the measurement results of electrochemical impedance spectroscopy (EIS), the pI of T24 lysate was about 6 ~ 7 because the impedance dropped after immunoassay in pH 7.0 buffer resulted from the negative charges within the protein. In addition, the impedance change before and after immunoassay increased when the pH value decreased. Consequently, the immunosensor with well-separated PANI nanofibers has demonstrated the feasibility of electrical sensing instead of fluorescent measurement, we believe this sensor has great potential of applications in biomedical detection.
摘要 I
ABSTRACT II
致謝 IV
目錄 V
圖目錄 VIII
符號彙編 XI
第一章 緒論 1
1.1 前言 1
1.2 文獻回顧 4
1.2.1 聚苯胺材料合成與應用 5
1.2.2 聚苯胺修飾電極於免疫分析 10
1.3 研究目的 12
1.4 研究架構與流程 13
第二章 電化學分析 16
2.1 電化學阻抗檢測 16
2.1.1 相角與複阻抗 17
2.1.2 電化學阻抗頻譜分析 18
2.1.3 阻抗頻譜圖與電路模型之關聯性 24
2.2 循環伏安法 26
第三章 實驗材料與方法 28
3.1 實驗材料與設備 28
3.1.1 實驗藥品 28
3.1.2 實驗設備 29
3.1.3 溶液配製 36
3.1.4 陽極氧化鋁模仁製備 37
3.1.5 聚苯胺奈米探針(纖維)製作 39
3.2 實驗架構與方法 41
3.2.1 實驗架構 41
3.2.2 免疫分析之流程 41
3.2.3 免疫分析之量測方法 42
第四章 結果與討論 45
4.1 聚苯胺奈米纖維之感測器製備 45
4.1.1 氧化鋁模板擴孔時間控制 45
4.1.2 質子酸對聚苯胺奈米探針形貌的影響 46
4.2 聚苯胺奈米探針之免疫分析 48
4.2.1 免疫反應之螢光響應 49
4.2.2 免疫反應之循環伏安法檢測 50
4.2.3 免疫分析之電化學阻抗分析 51
第五章 結論與未來展望 63
5.1 結論 63
5.2 未來展望 64
參考文獻 65
作者簡介 67
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