應用表面電漿調控螢光生命週期及螢光各向異性之生醫感測器研究

Abstract

本論文探討表面電漿共振現象對於螢光生命週期以及螢光各向異性之調變，以及未來應用於生醫感測器之方法。螢光生命週期與螢光各向異性訊號具有定性以及定量的分子資訊；相較於螢光光強訊號，更適合應用於臨床應用以及活體量測時的複雜系統。表面電漿共振現象對於螢光訊號之調變，將有利於螢光訊號分析。本研究中使用螢光標定之去氧核醣核酸，進行金奈米粒子之自組裝製程。藉由直徑為二十奈米之金奈米粒子所產生的局域性表面電漿共振現象，同時，利用去氧核醣核酸長度，決定螢光分子與金奈米球之間的距離，以控制螢光分子之螢光生命週期。另一方面，生醫感測器必須具有微型化、可攜式之特點。因此，研究中螢光訊號之量測使用發光二極體作為光源，而樣品產生之螢光則分別由增強型電荷耦合器、與光電倍增管的架構偵測。實驗結果中，驗證螢光分子確實受到局域性表面電漿共振的影響；當螢光分子與金奈米粒子兩者之間距離越近時，螢光分子產生光強變弱以及螢光生命週期變短的現象，證明螢光生命週期能夠藉由控制螢光分子和金奈米粒子之間的距離以進行調變。最後，實驗中驗證螢光生命週期之變化，亦影響螢光各向異性訊號。本論文的研究成果提供生醫檢測時之分子量測的發展方向。

In this thesis, I investigated the modulation of fluorescence lifetime and anisotropy by localized surface plasmon resonance (LSPR), with the goal for biosensor applications. Fluorescence lifetime and anisotropy provide qualitative and quantitative information on molecular states. Compared with fluorescence intensity, fluorescence lifetime and anisotropy are better for clinical application and in vivo measurements of complex bio-system. I utilized deoxyribonucleic acid (DNA) with Alexa 532 labelling, and then fabricated DNA-directed self-assembly of gold nanoparticle. Gold nanoparticles of 20 nm in diameter was chosen to make LSPR device. DNA was employed to exactly control the distance between dye molecule and gold nanoparticle, in order to control fluorescence lifetimes of Alexa 532. To realize biosensor with miniaturized size and portability, light emitting diode (LED) was used as the light source to excite the fluorescent specimen. In my experiments, fluorescence lifetime and anisotropy signals were detected by intensified charge-coupled device (ICCD) and photomultiplier (PMT), respectively. The results in this thesis demonstrated that fluorescence signals were clearly affected by LSPR. As the distance between the flourophore and gold nanoparticle decreased, the fluorescence intensity became quenched and lifetime was decreased. This phenomenon showed that fluorescence lifetime of dye molecule was controllable, simply by changing the distance between dye molecule and gold nanoparticle. I also demonstrated that fluorescence anisotropy was tunable by changing fluorescence lifetime of fluorophore. The results of this thesis provides a method to design biosensor for molecule detection.