近年來，胞外體（EVs）在生物學領域引起了高度關注。隨著分離和分析技術的顯著發展，越來越多胞外體的生理和病理功能被發現。胞外體在細胞通訊中擔任重要的角色，它們有助於將特定的脂質、蛋白質和核酸從親代細胞轉移到受體細胞。由於胞外體的多種特性，它們可以應用於生物領域的臨床醫學，也得以作為藥物載體。為了更深入的研究胞外體，我們應優先開發一種支持標準化且有效分析胞外體的方法。
本研究採用熱透鏡顯微鏡（TLM）結合微流體晶片進行胞外體量化分析。熱透鏡顯微鏡是一種靈敏的檢測技術，適用於檢測非螢光和無標記分子。通過與微流體晶片結合，能夠顯著增強樣品的特異性，也可以嘗試操控樣品。然而，熱透鏡顯微鏡在微流體領域仍未得到充分利用。為了擴大其用途並開發一種有效的分析方法，本研究主要集中在利用熱透鏡顯微鏡搭配微流體晶片對胞外體量化分析。

Recently, extracellular vesicles (EVs) raised high attention in the biological field. Since the separation and analysis technologies have been significantly developed, more and more physiological and pathological functions of EVs were discovered. EVs were found to play an important role in intercellular communication; they help transfer specific lipids, proteins, and nucleic acids from parent cells to recipient cells. Due to the variable characteristics of EVs, they can be applied to clinical medicine in the biological field, and also can be potentially served as drug carriers. In order to investigate more about EVs, a standardized and effective approach supporting EV analysis should be well-developed in priority.
In this research, the desk-top thermal lens microscope (DT-TLM) combined with a microfluidic chip was adopted to perform EV quantification analysis. Thermal lens microscopy (TLM) is a sensitive detection technique that is suitable for detecting non-fluorescent and label-free molecules. By integrating with the microfluidic chip, the sample specificity could be enhanced and the sample can also be possibly manipulated. However, TLM still remains underutilized within the field of microfluidics. To broaden its use and to develop an effective analysis approach, this research mainly focused on utilizing TLM as a quantification technique for EVs in a well-designed microfluidic chip.

LIST OF FIGURES VI
LIST OF TABLES VIII
CHAPTER1: Introduction 1
1.1 Research background 1
1.2 Introduction of Extracellular Vesicles (EVs) 1
1.3 EVs quantification techniques 4
1.3.1 Optical microscopy 5
1.3.2 Raman spectroscopy 6
1.3.3 Dynamic light scattering (DLS) 8
1.3.4 Nanoparticle tracing analysis (NTA) 9
1.3.5 Tunable resistive pulse sensing (tRPS) 11
1.3.6 High-resolution flow cytometry (hFC) 13
1.4 Introduction of Thermal Lens Microscopy (TLM) 15
1.4.1 Working principle of TLM 16
1.4.2 Instrument 23
1.4.3 Applications 24
1.5 Objectives 25
CHAPTER2: Experimental design and methods 26
2.1 Flow chart 26
2.2 Apparatus and experimental setup 27
2.2.1 Apparatus 27
2.2.2 Microfluidic device 28
2.2.3 Experimental setup 29
2.3 Experiment design 31
2.3.1 Concentration determination 34
2.3.2 Individual particle detection 35
2.4 Experiment methods 36
2.4.1 Sample preparation 36
2.4.2 Tunable resistive pulse sensing 38
2.4.3. Absorbance measurement 39
2.4.4 Concentration determination 40
2.4.5 Individual particle detection 41
CHAPTER 3: Results and discussion 43
3.1 Preliminary experiment 43
3.1.1 Absorption spectrum 43
3.2 Concentration determination 48
3.2.1 Measurement of sunset yellow FCF 48
3.2.2 Measurement of MCF-7 EVs 50
3.3 Individual particle detection 55
3.3.1 Verification of detection 55
3.3.2 Standard of determining the suitable concentration 56
3.3.3 Effect of flow rate on individual particle detection 56
3.3.4 Calibration curve of standard dyed polystyrene nanoparticles 58
3.3.5 Measurement of HEK-293T EVs 59
3.3.6 Influence of using different time constants 60
3.3.7 Peak profile of dyed polystyrene nanoparticles and HEK-293T EVs 61
CHAPTER4: Conclusion and future work 63
REFERENCES 64

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