核酸適合體式光學波導共振生物感測器於凝血酶之檢測

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林聖富(Sheng-Fu Lin) 查詢紙本館藏

畢業系所

光電科學與工程學系

論文名稱

核酸適合體式光學波導共振生物感測器於凝血酶之檢測
(Application of Aptasensor by Using Guided Mode Resonance for Thrombin Detection)

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摘要(中)

本研究在研發一新型核酸適合體式生物感應器（aptasensor）。隨著科技的進步，衛生醫療保健的觀念越來越受到大家的注意，其中預防醫學的觀念也越來越普及。檢測微量生物指標（biomarker）有助於疾病的早期發現與早期治療，而生物感測器在此扮演很重要的角色。除了疾病的檢測之外，更可將檢測器用於監控環境或戰場運用，固有必要發展生物感測器。本研究之感測器結合光學波導共振元件與核酸適合體。波導共振元件（guided mode resonance device）做為生物感測器中的訊號換能元件（transducer），而核酸適合體（aptamer）為辨識元件（recognize element）。核酸適合體於生物檢測中有需多的優點，除了對檢測物有專一的鍵結能力（specific binding），此分子為人工合成之序列、研發期相對於一般抗體短、易於修飾且不影響與待測物的親和力、對應之檢測物廣與保存容易等優點。綜合以上辨識元件的優點再搭配上波導共振元件在生物檢測上的優勢，這將成為深具潛力的生物感測器。
於本研究中我們已經成功的將核酸適合體固定於波導共振元件之表面，並且將檢測系統雜訊降低到 0.006 nm 的波長偏移量，等效檢測極限為 8.57*10-5 的折射率變化。於實驗中成功即時檢測不同濃度之凝血酶，檢測範圍從 0.25 microM 到1.5 microM 之濃度與其訊號飄移量呈線性關係。從即時檢測的資料中可提供動力學資訊，親和力常數（affinity constant）Ka 皆介於 10-6 的等級，最後此生物晶片達到可重複性使用與重現性的目標。

摘要(英)

In this thesis we develop a novel biosensor which combines aptamer and guided mode resonance device which is called aptasensor. As the technology improving, the life quality of humankind also improves, and people takes more concerned with the hygiene. In this generation, preventive medicine is popularizing, and many studies show some disease or cancer which will release some special biomarker before onset, so biosensors play an important role here. If we could detect the trace biomarker early, we have more opportunity to cure the diseases. Besides, biosensors are also applied to health care, monitoring environment, biomedical research or battle field. So developing biosensor is very important.
Basically biosensors have two parts, one is the recognizing element and the other is the transducer. In this thesis we use the aptamer to be the recognizing part (immobilize molecular) and optical guided mode resonance (GMR) device to be the transducer. Aptamer have high specific binding to ligands, and aptamer is a DNA or RNA fragment which can be synthesized in vitro. According to GMR’s special and designable sensitivity to surface detect and many potential advantages, so we apply it to biosensor.
In the long run of this research, we achieve the thrombin sensing in concentration from 0.25 microM to 1.5 microM, the noise in our system is around 0.06 nm, limit of detection (LOD) is 8.57*10-5 RIU. Finally the kinetics from our real time experiment results is gotten, the affinity constant (Ka) is around order.

關鍵字(中)

★ 凝血酶
★ 生物感測器
★ 核酸適合體
★ 波導共振

關鍵字(英)

★ biosensor
★ aptamer
★ guided mode resonance
★ thrombin

論文目次

論文電子檔授權書 ..................................................................................................................... i
論文指導教授推薦書 ................................................................................................................ ii
論文口試委員審定書 ............................................................................................................... iii
摘要 .......................................................................................................................................... iv
Abstract ..................................................................................................................................... vi
致謝 ......................................................................................................................................... vii
目錄 ........................................................................................................................................ viii
圖目錄 ....................................................................................................................................... x
表目錄 ..................................................................................................................................... xii
第一章序論 ............................................................................................................................ 1
1-1 生物感測器 ................................................................................................................ 2
1-2 光學式波導共振元件 Guided mode resonance 應用於生物感測.......................... 5
1-3 核酸適合體 Aptamer 應用於生物感測 .................................................................. 8
1-4 凝血酶簡介 .............................................................................................................. 13
1-5 抗凝血酶核酸適合體 Thrombin binding aptamer ................................................. 15
1-6 研究動機 .................................................................................................................. 16
第二章波導共振元件 Guided mode resonance device ................................................... 17
2-1 波導共振元件理論 .................................................................................................. 18
2-1.1 光波導原理 ........................................................................................................ 18
2-1.2 繞射光柵原理 .................................................................................................... 20
2-1.3 波導共振基本原理 ............................................................................................ 22
2-2 波導共振元件之設計與模擬 .................................................................................. 25
第三章製程、實驗與量測 .................................................................................................. 32
3-1 波導共振元件製作與微流道系統 .......................................................................... 33
3-1.1 波導共振元件之製作 ........................................................................................ 33
3-1.2 微流道系統： .................................................................................................... 34
3-2 實驗儀器與藥品 ...................................................................................................... 37
3-2.1 晶片製作： ........................................................................................................ 37
3-2.2 表面改質： ........................................................................................................ 38
3-2.3 後端量測： ........................................................................................................ 39
3-3 實驗步驟 .................................................................................................................. 41
3-3.1 表面改質： ........................................................................................................ 41
3-3.2 流體即時檢測： ................................................................................................ 43
第四章實驗結果與討論 ...................................................................................................... 45
4-1 晶片製作 .................................................................................................................. 46
4-2 表面改質 .................................................................................................................. 51
4-3 酒精實驗 .................................................................................................................. 54
4-4 凝血酶檢測實驗 ...................................................................................................... 55
4-5 討論 .......................................................................................................................... 65
第五章結論與未來展望 ...................................................................................................... 70
5-1 結論 .......................................................................................................................... 71
5-2 未來展望 .................................................................................................................. 72
第六章參考文獻 .................................................................................................................. 73

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指導教授

張正陽(Jenq-Yang Chang)

審核日期

2009-7-23

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