應用生命週期技術進行螢光與激發光分離

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姓名

楊皓亘(Hao-Xuan Yang) 查詢紙本館藏

畢業系所

光電科學與工程學系

論文名稱

應用生命週期技術進行螢光與激發光分離
(Unmixing of fluorescence and excited light using fluorescence lifetime imaging microscopy)

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至系統瀏覽論文 (2027-6-7以後開放)

摘要(中)

拉曼光譜經常會伴隨著大量的螢光干擾，為了減少螢光干擾，我們從螢光能量共振轉移技術(fluorescence resonance energy transfer, FRET)看到了解決方法，利用螢光生命週期的特性，將螢光與拉曼訊號進行分離，由於目前只是初步的系統開發，我們使用生命週期特性與拉曼光譜相似的激發光代替拉曼訊號進行量測，確認系統是否能準確分離螢光與激發光，後續便能拓展回拉曼訊號與螢光的分離。我們使用頻率域的螢光生命週期技術(FLIM)進行實驗，由於頻率域需要對於光源和偵測訊號進行弦波調制，我們會先進行電路設計，設計出可以調制雷射二極體的驅動電路，後續使用零差檢測的方法，使用類比乘法器將偵測訊號與調制雷射的參考訊號相乘，便能透過此系統量測其等效生命週期，將螢光和激發光進行線性分離，得到激發光與螢光的強度比。

摘要(英)

Raman spectroscopy is often accompanied by a large amount of fluorescence. In order to reduce background fluorescence, we have seen a solution through Förster resonance energy transfer (FRET) technology, using the characteristics of the fluorescence lifetime, the fluorescence and raman signal can be separated. Since it is only a preliminary system development, we use excitation light with similar lifetime characteristics to Raman spectrum instead of Raman signal for measurement to confirm whether the system can accurately separate fluorescence and excitation light. Follow-up This can be extended back to the separation of Raman signal and fluorescence. We use the fluorescence lifetime microscopy (FLIM) in the frequency domain for experiments. Since the frequency domain requires sine wave modulation for the light source and detection signal, we will first design the circuit and design a driving circuit that can modulate the laser diode. Use the homodyne detection method to multiply the detection signal and the reference signal of the modulated laser by an analog multiplier, and then the equivalent lifetime can be measured through this system, and the fluorescence and excitation light can be separated. The intensity ratio of excitation light and fluorescence can be obtained.

關鍵字(中)

★ 生命週期
★ 螢光分離

關鍵字(英)

論文目次

中文摘要 i
Abstract ii
致謝 iii
目錄 iv
圖目錄 vi
表目錄 viii
第一章緒論 1
1.1 研究動機 1
1.2 拉曼光譜螢光抑制技術 2
1.3 螢光顯微技術介紹 4
1.4 螢光生命週期技術(FLIM)介紹 6
1.5 研究目標 8
第二章實驗原理 10
2.1 螢光生命週期量測 10
2.2 生命週期極座標分析法 12
2.3 極座標之線性分離訊號法 14
第三章實驗方法 16
3.1 系統架構 16
3.2 程式控制 20
3.3 雷射二極體驅動電路製作 21
3.4 系統校正 24
3.5 樣本製備及分析方法－羅丹明6G (R6G)、伊紅 Y (Eosin Y) 26
第四章實驗結果 28
4.1 雷射與系統的調制深度分析 28
4.2 光電倍增管PMT校正平面量測 30
4.3 系統驗證及分析 33
4.4 以生命週期技術分離螢光及激發光 34
第五章結論與展望 42
參考文獻 43

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指導教授

陳思妤(Szu-Yu Chen)

審核日期

2022-7-26

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