表面增強拉曼散射探針及微流道系統 用於癌症細胞及外泌體表面生物標記物的多重檢測;Immuno-functionalized Surface-Enhanced Raman Spectroscopy-Probes (SERS-Probes) and Microfluidic System for Multiplexed Detection of Cell and Exosome Surface Biomarkers in Cancer Diagnosis

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Title:	表面增強拉曼散射探針及微流道系統用於癌症細胞及外泌體表面生物標記物的多重檢測;Immuno-functionalized Surface-Enhanced Raman Spectroscopy-Probes (SERS-Probes) and Microfluidic System for Multiplexed Detection of Cell and Exosome Surface Biomarkers in Cancer Diagnosis
Authors:	闕華恩;Chueh, Hua-En
Contributors:	生醫科學與工程學系
Keywords:	表面增強拉曼探針;表面生物標記物;拉曼多重檢測;最小平方法;微流道;熱壓黏合法;COMSOL模擬;Immuno-functionalized SERS-probes;Surface Biomarkers;SERS Multiplexed Assay;Classical Least Square;Microfluidics;Thermal Bonding;COMSOL simulation
Date:	2023-08-21
Issue Date:	2024-09-19 15:20:51 (UTC+8)
Publisher:	國立中央大學
Abstract:	癌症的早期檢測有助於在疾病最前期識別出癌症或癌前病變，這可以提高存活率及減少發病率。過去十年中，使用體液進行癌症相關分析的液態檢體引起了廣泛的關注。體液中含有很多生物分子，並有可能代表疾病狀態，且檢體方便取得，所以是一個新興的疾病檢測方法。隨著研究的成熟，許多研究者發現，使用多個生物標記物而不僅是單一生物標記物是實現此目標更可行的方法。因此，在液態檢體中檢測細胞或外泌體上的不同膜蛋白標記物，在癌症診斷中具有重要意義。目前多重檢測最常用的儀器是流式細胞儀，其面臨的問題是螢光染劑的散射光帶寬重疊，無法有效區分不同檢測物，而拉曼光譜有著很窄的帶寬，可以透過光譜解析以獲得不同成分的組成。此外，微流道系統是處理少量液態檢體(如血液)的好工具，但傳統的製造方法(如曝光顯影技術)耗時耗財。此研究合成出不同組免疫功能化的奈米等級表面增強拉曼探針（SERS-probes）來檢測多個目標分子，且使用雷射雕刻和熱壓黏合方法快速製作微流道試片，建立一套癌症生物標記物的檢測系統。另外，使用COMSOL模擬可以優化微流道系統的設計，而將最小平方法用在化學成分推論，則可以進行拉曼光譜解析以獲得不同成分的組成。再者，由於乳癌的廣泛流行且該領域已有相當深入的研究，我們先選擇其進行概念驗證。本研究著眼於拉曼多重檢測的拉曼分子選擇方式、表面增強拉曼探針合成步驟的探討、及簡易的微流道製程方法，並且提出了一種結合免疫功能化的表面增強拉曼探針和微流道技術的方法，為細胞和外泌體表面生物標記物的多重檢測建立了有前景的平台。這項研究有助於開發透過液態檢體進行早期癌症檢測的新型診斷工具，並為未來的研究者提供適合更深入研究的方向。;Liquid biopsies have the ability to reveal the presence or absence of illness. It is especially suitable for early cancer detection because of the easy availability of body fluids. Furthermore, many researchers concluded that using multiple biomarkers instead of one is more feasible. As a result, identifying different membrane proteins as biomarkers on cells or exosomes in liquid biopsy has gained importance in cancer diagnosis. Currently, the most common instrument for multiplexing is flow cytometer. One of its challenges is fluorescence overlapping, making it difficult to effectively distinguish different analytes. On the other hand, Raman spectroscopy features a narrow bandwidth, various components can be differentiate through spectral analysis. Moreover, microfluidics are good tools for handling small quantities of liquid samples, but traditional fabrication methods like soft lithography are time-consuming and costly. This study employed laser cutting and thermal bonding method to quickly fabricate microfluidic chips and also successfully synthesized different immuno-functionalized surface-enhanced Raman scattering-probes (SERS-probes) to detect multiple target molecules at the same time, taking advantage of its narrow bandwidth compared to fluorescence. Also, COMSOL simulation is used to optimize microfluidics designs, while classical least square demultiplexs SERS spectra. Additionally, Breast cancer is chosen as the primary demonstration for this study due to its wide prevalence and the in-depth research conducted in this field. Our results proposed an integrated method of immuno-functionalized SERS-probes combined with microfluidics to provide a promising platform for the multiplex detection of cell and exosome surface biomarkers. This research potentially contributes to developing new diagnostic tools for the early detection and also point a direction for future research.
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