吾人研究羅丹明B染料與微米金板之間的距離對於金屬增強螢光效應的影響，使用抗生物素蛋白-生物素複合物(Avidin-biotin complex)作為間隙物連接染料分子與微奈米基質，利用螢光生命期影像顯微技術選擇單一樣品並擷取染料螢光生命期獲得螢光放光強度的影響。透過合成70 nm的奈米金球與0.6-1.0 m的微米金板當基質，並以逐層(Layer-by-layer)法將抗生物素蛋白及生物素堆疊其上。改變此間隙物之厚度。所收取到的螢光生命期均呈現雙自然指數衰減，在4-8 nm厚度中較短的生命期18-23 ps，較長的生命期於微米金板上依一層至四層抗生物素蛋白-生物素雙層為249、271、304和348 ps生命期隨厚度增加而變長，在間隙物四層時，距離約8 nm有最大的光子計數，相當於單層時的約四倍。我們推導動力學模型以適解此動力行為，在奈米金球為基質時，只有單層抗生物素蛋白，受激發的染料易將以表面電漿共振能量傳遞至高偶極模式(High-dipole modes)，因為高偶極模式之金無法放光，使得能量淬息比例高且放光弱。在微米金板時，若染料與金屬表面相距約5 nm時，即含1-2層抗生物素蛋白，染料傳遞至暗模式仍高。隨著間隙物厚度增加，激發態的染料分子受金電場加強放光速率與傳遞至暗模式速率相當，使得螢光增強效果顯示出來，因此染料於四層抗生物素蛋白上約8 nm厚度可看看到最高的光子計數。

We study the effects of metal-enhanced fluorescence (MEF) on Rhodamine B dye molecule by gold microplates. The avidin-biotin complex was used as a spacer to connect Rhodamine B and the microplate. Fluorescence lifetime image microscopy (FLIM) combined with time-correlated single-photon counting (TCSPC) was used to detect the emission image and the lifetime of dye on single particle. Spherical gold particles dia. 76 nm and gold hexagon plates size 0.6-1.0 m were synthesized and the avidin-biotin complexes were placed layer-by-layer up to 4 layers onto the microplates. For all emission curves we observed biexponetial decay with the first lifetime 18-23 ps and the second lifetime 249, 271, 304, and 348 ps for one to four avidin-biotin bilayers on gold plates, respectively. Our results reveal that the spacer with four bilayers (8 nm) has the most emission intensity counts. We derive a kinetic model to explain the biexponential behavior. In the case of gold nanospheres, the excited dye transferred its energy mostly to the high order modes of gold nanospheres which are nonemissive. Hence, the quenching is dominant. At gold microplates, when the distance between the dye and the gold surface is less than 5 nm, similar results are obtained as those in the nanospheres. While the thickness of spacer increases, the rate of enhancement by the gold plates during plasmonic excitation is comparable with the energy quenching rate. The process of the enhanced emission via excitation becomes dominate to show increased photon counts in the FLIM measurements.

目錄
第一章 序論 1
1.1螢光增強效益簡介 1
1.2研究動機 2
1.3研究方法 2
第二章 基本原理 5
2.1金屬表面電漿子現象 5
2.2表面電漿共振衰減機制 9
2.3當染料與金屬奈米粒子共存時的局部電場增強效應 11
2.4金屬增強效應的動力學機制 11
第三章 文獻回顧 21
3.1金屬奈米粒子增強螢光效應 21
3.1.1 影響因素 21
3.1.1.1距離因素 21
3.1.1.2染料吸收及放光與表面電漿共振吸收譜帶重疊 25
3.1.1.3金屬粒徑 30
3.2金屬奈米粒子淬息螢光效應 31
3.3抗生物素蛋白的特性 34
第四章 樣品製備 37
4.1微米金板 37
4.2奈米金球 37
4.3製備FLIM樣品 38
4.3.1 單層抗生物素蛋白於奈米金球/微米金板 39
4.3.2 雙層抗生物素蛋白於微米金板 40
4.3.3 三層抗生物素蛋白於微米金板 40
4.3.4 四層抗生物素蛋白於微米金板 41
第五章 儀器架設 42
5.1時間相關單光子技術系統 42
5.1.1 原理 42
5.1.2 電子元件 45
5.1.2.1分數式時間鑑別器 45
5.1.2.2時間-振幅轉換器 47
5.1.2.3類比數位轉換器 48
5.2共軛焦螢光生命期影像顯微鏡 48
5.2.1 原理 48
5.2.2 儀器架設 49
5.2.2.1雷射系統 50
5.2.2.2壓電平移平台 51
5.2.2.3單光子雪崩二極體 51
5.2.2.4時間相關單光子計數 52
5.2.2.5SymPhoTime軟體 52
5.3其他實驗系統 52
5.3.1 紫外-可見光吸收光譜儀(U-3900H Specttrophotometer, Hitachi) 52
5.3.2 靜態螢光光譜(F-7000 FL Spectrophotometer, Hitachi) 53
5.3.3 掃描式電子顯微鏡(Field Emission Scanning Eletron Microscopy, FESEM) 53
第六章 實驗結果與討論 54
6.1製備樣品之特性 54
6.1.1奈米金球與微米金板之SEM圖 54
6.1.2奈米金球與微米金板之靜態吸收光譜 55
6.1.3染料之靜態吸收與螢光光譜 57
6.2FLIM影像之校正 58
6.3FLIM樣品之時間解析光譜 61
6.3.1單層抗生物素蛋白於奈米金球之時間解析螢光光譜 61
6.3.2單層抗生物素蛋白於微米金板之時間解析螢光光譜 63
6.3.3雙層抗生物素蛋白於微米金板之時間解析螢光光譜 66
6.3.4三層抗生物素蛋白於微米金板之時間解析螢光光譜 69
6.3.5四層抗生物素蛋白於微米金板之時間解析螢光光譜 72
6.4分子動力學模型 76
6.4.1奈米金球增強螢光效應之動力學模型研究 78
6.4.2微米金板增強螢光效應之動力學模型研究 79
第七章 結論 84
參考資料 85

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