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研究生:余聲宏
研究生(外文):Sheng-Hong Yu
論文名稱:光纖漸逝波感測器應用於微量生物分子檢測
論文名稱(外文):Fiber Optics Evanescent Wave Biosensor for Biomolecular Detection
指導教授:江惠華江惠華引用關係
指導教授(外文):Huihua Kenney Chiang
學位類別:碩士
校院名稱:國立陽明大學
系所名稱:醫學工程研究所
學門:工程學門
學類:生醫工程學類
論文種類:學術論文
論文出版年:2004
畢業學年度:92
語文別:中文
論文頁數:116
中文關鍵詞:光纖漸逝波三明治螢光免疫分析法結合常數解離常數
外文關鍵詞:Fiber opticEvanescent wavesandwich immunofluorescence assayassociation constantdissociation constant
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  • 被引用被引用:12
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光纖漸逝波生物感測器具有低成本、易於設計且即時、即位動態監測生物分子間交互作用等功能,目前已廣泛應用於臨床生物醫學、試藥開發、國防工業以及環境生化監測上。研究中採用稀釋氫氟酸蝕刻液,製備核心直徑為400μm、檢測區域長度為5cm之漸逝波式二氧化矽玻璃光纖感測器。同時利用胺基矽烷化合物修飾無機光纖表面,藉由三明治螢光免疫分析法,量測漸逝波場內Alexa Fluor 488螢光指示劑,結合相對濃度待測生物分子複合物之含量。
本研究目前已完成光纖漸逝波量測系統之建置。在實驗架構上,選擇於光纖之側向接收不同濃度螢光分子,其與待測分析物結合之相對受激散射螢光訊號,並於光纖末端鄰接反射率為95%之面鏡,以便有效利用激發光源。實驗結果顯示,光纖經由化學蝕刻液去除纖衣層後,由於表層極為平整,故可獲致均勻度較佳之漸逝波激發場。目前實驗已能精確控制光纖之蝕刻深度,於室溫條件下為0.1046μm/min。
在系統靈敏度方面,針對FITC純螢光分子檢測,目前最低可測濃度達1μg/mL,對於IgG–Anti rabbit IgG檢測,目前可達100ng/mL,對於Biotin– Streptavidin生物分子量測,目前可達50ng/mL,同時實驗反應腔之最低樣本體積含量僅需500μL。於化學動力學參數計算上,我們評估10分鐘內的即時螢光訊號反應強度,初步推估IgG–Anti rabbit IgG之結合常數、解離常數以及親合力常數分別為2.09×105M-1s-1、1.63×10-3 s-1、1.29×108 M-1,Biotin–Streptavidin之結合常數、解離常數以及親合力常數分別為2.06×105M-1s-1與3.93×10-3s-1、5.24×107 M-1,此結果與目前商用檢測產品之典型值範圍相符,預期此一系統未來將可應用於臨床的活體生物樣本檢測上。
The evanescent wave fiber optic biosensors (EWFOBS) have the advantages of low cost, compact design and real-time in-situ monitoring for bio-molecular interaction. In this study, we utilize the BOE solution (dilute hydyofluorid acid) to etch the 400 μm, 5cm active region silica optical fibers, and to modify the surface of optical fiber with 3-aminopropyltriethoxysilane (APTS). In the evanescent field adjacent to the surface of fiber, fluorescent indicators Alexa Fluor 488 that is related to the concentration of the target analytes are detected by the structure in sandwich immunofluorescence assay.
We propose and set up a EWFOBS system. The different concentration of analytes combine with fluorescent dyes conjugation, proportional to scattering fluorescence signals on the flank of the fiber are measured. In order to achieve the multi-excitation effectively, the 95% aluminum coating mirror is placed to near the end of the fiber. Preliminary experiments have shown that the surface of the fiber are very flatness, it means that the evanescent wave is very uniform onto the surface, and we can control the etching depth exactly. The etching rate is 0.1046μm/min now.
The sensitivity in estimation is 1μg/mL for FITC, 100ng/mL for IgG–Anti rabbit IgG and 50ng/mL for Biotin–Streptavidin respectively. The aqueous sample in all detection is 500μL now. The chemical kinetics calculations in this thesis are assessed in the time slot of 10 minutes, and we can obtain the association constant (ka), dissociation constant (kd) and affinity constant (K) are 2.09×105M-1s-1, 1.63×10-3 s-1 and 1.29×108 M-1 respectively for IgG–Anti rabbit IgG, and 2.06×105M-1s-1, 3.93×10-3 s-1 and 5.24×108 M-1 respectively for Biotin-Streptavidin. The parameters of kinetics are fit the typical range of commercial products, and we can expect this system to be capable of applying on in-vivo clinical detection in near future.
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