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系統識別號 U0026-0812200911181353
論文名稱(中文) 應用電性鑑別偵測法於微免疫分析晶片之研究
論文名稱(英文) Study of Micro-Chip for Electro-Immunoassay in Real-Time
校院名稱 成功大學
系所名稱(中) 工程科學系碩博士班
系所名稱(英) Department of Engineering Science
學年度 92
學期 2
出版年 93
研究生(中文) 蘇凱群
研究生(英文) Kai-Chun Su
學號 n9691151
學位類別 碩士
語文別 中文
論文頁數 125頁
口試委員 指導教授-林裕城
口試委員-陳顯禎
口試委員-林弘萍
中文關鍵字 奈米粒子  電性鑑別  酵素免疫分析  生物晶片  微機電 
英文關鍵字 nano-particle  ELISA  Electro-Immunoassay  Biochip  MEMS 
學科別分類
中文摘要   本研究目地為發展電性鑑別法於微量化的酵素免疫分析檢驗,並致力於提昇檢疫的各項反應表現。運用微機電系統 (MEMS, Micro-Electro-Mechanical Systems)的製程技術來設計與製造『微電極免疫分析檢驗晶片』,以晶片微小化達到容納大量資訊、減少檢測時間、降低試劑成本及提高檢測靈敏度等檢疫要求。
  電性免疫檢測法(Electro-Immunoassay)運用電磁學中的集膚效應作為理論依據,有別於傳統呈色判定的酵素免疫分析,並利用高頻電子訊號方式來完成實驗鑑定與定量工作。鑑別方式藉由相位、阻抗、電阻、電抗於頻率響應後的變化結果,來完成免疫檢驗的分析與判斷。此外,依循物理機制的設計概念,研究將晶片搭配金奈米粒子(13 nm)作兩個不同方面的探討:(一)晶片表面先接著奈米粒子以提高檢測區內的反應表面積,是否能有效放大電性訊號的反應結果;(二)在檢疫反應最後加入帶有結合奈米粒子的二次抗體以捕捉檢測抗原,是否能造成晶片表面的阻抗變化。以上討論均以運用奈米粒子的微觀物理效應而達到快速鑑別的目地。
  綜合實驗結果發現,電性偵測在頻域上具有明顯的鑑別性,而且抗體與抗原的即時反應過程可表現於動態的電性量測上,而在檢測極限方面,待測抗原濃度的偵測靈敏度可達到 10-8 g/ml。此外,金奈米粒子的運用雖無助於偵測靈敏度的提升,但有助於量測結果的判斷與鑑別。由此可知,電性鑑別方式可運用於酵素免疫分析的檢疫反應上。
英文摘要   In this study, a novel Micro-Electrode Biochip for Immunoassay was designed and fabricated by using MEMS (Micro Electro-Mechanical Systems) technique. The characteristics of Micro-Electrode Biochip with a capacity of efficient informative nano-samples had been developed for the purpose of decreasing reaction time and the cost. On the Biochip, we were devoted to the study of the sensitivity of detection limit and took advantages of traditional ELISA in time.
  Following the skin effect from Maxwell equation, we analyzed the signals from the electrodes on the chip. Measured impedance changes in frequency domain would help us to identify the result of Electro-Immunoassay. In order to improve the reaction and economize on time, conjugated with Au nano-particles was adopted to amplify the output signal of impedance measurement and distinguish the specific antigen easily and rapidly. Four parameters, phase, impedance, resistance and reactance, of electric properties under different testing voltages had been discussed to identify in the analysis of frequency/impedance spectroscopy.
  Finally, we had found out some interest results in characteristic curves of electric signals. The reaction between antibody and antigen in real time could be detected in electrical measurement through high frequency domain. Furthermore, the detection limit in the concentration of antigen was 10-8 g per milliliter. In another hand, Micro-Electrode Biochip conjugated with Nano-Au (13 nm) for Electro-Immunoassay was conveniently in signal discrimination. The results of this study had been confirmed excellent influences in Immuno-topic.
論文目次 中文摘要 I
Abstract II
誌謝 III
目錄 IV
表目錄 XI
第一章 前言 1
1-1 研究背景與動機 1
1-2 文獻回顧 4
1-3 研究目標 7
1-4 實驗架構 8
1-5 論文架構 9
第二章 電性偵測應用於蛋白質酵素免疫分析之原理 10
2-1 酵素免疫分析簡述 11
2-2 電性偵測的實驗模型建立與方法 14
2-3 微電極晶片的表面集膚效應 17
2-4 微電極晶片於電解質溶液的離子傳遞效應 20
第三章 電性偵測平台系統介紹 23
3-1 阻抗頻譜分析儀說明 23
3-2 電性鑑別分析系統的建立與操作準備步驟 26
3-3 實驗的相關參數設計 29
第四章 晶片設計與製作 32
4-1 晶片結構與基材選擇 32
4-2 光罩與設計 35
4-3 晶片製程 38
4-3-1 基板清洗 39
4-3-2 金屬蒸鍍 42
4-3-3 微影 45
4-3-4 金屬蝕刻 50
4-3-5 PDMS固化 54
4-3-6 晶片滅菌 58
第五章 晶片測試與實驗 59
5-1 微電極免疫分析晶片 59
5-1-1 基材選擇與晶片製程 60
5-1-2 微電極晶片的電性量測結果 63
5-2 增加反應表面積的實驗 68
5-3 改變晶片導電度的實驗 70
5-4 即時偵測的實驗 73
第六章 結果與討論 75
6-1 反應表面積增加的實驗結果 75
6-1-1 奈米粒子之高表面積特性對接著比例的影響 75
6-1-2 干擾與雜訊的分析探討 79
6-1-3 增加反應表面積的可行性 83
6-2 晶片表面導電度改變的實驗結果 84
6-2-1 抗體的阻抗電性反應 87
6-2-2 抗原的阻抗電性反應 92
6-2-3 電性鑑別的專一性 97
6-2-4 偵測的靈敏度 102
6-2-5 乾式量測與濕式量測的運用 107
6-3 即時偵測 112
第七章 結論與建議 114
7-1 結論 114
7-2 建議 117
7-3 未來展望 119
附錄 120
參考資料 121

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