2008-08-012024-05-17https://scholars.lib.ntu.edu.tw/handle/123456789/692868摘要：本計劃主要目的主要根據前期計畫「應用奈米粒子布朗運動行為開發生物分子感測晶片之研究」進一步開發生物分子感測技術，本研究藉由第一年所發展奈米粒子鍵結抗體之技術，除了應用於布朗運動之生物分子感測技術外，也將此技術用於鍵結一般之蛋白質分子，生物蛋白分子之表面具有電荷，此外生物蛋白分子亦具有被電場極化之性質，藉由本實驗室過去所發展之界電泳(DEP)技術針對奈米粒子上是否鍵結蛋白質分子進行分離，由於懸浮奈米粒子與溶液中蛋白質分子之鍵結反應速率相對於壁面鍵結要高，因此可以達到快速反應的效果，此外，該技術亦可用於檢測濃度較低之蛋白質或抗原，藉此達到精密生物檢測之效果，未來可望成為其他生物巨分子或奈米檢測系統之應用基礎。 目前本實驗室已具備完整介電泳晶片開發之技術，並已針對血液等生物容易完成多項研究成果。目前已掌握微機電製程技術，並將往奈米製程技術發展。本計畫將以數百奈米粒徑之的奈米螢光膠狀粒子為媒介，鍵結生物液體中之蛋白質如血蛋白或球蛋白，並根據介電泳效應判定奈米粒子鍵結蛋白質前後之差異，並搭配全不同之電極設計已達到有效的分離效果，實驗中將以高倍率之螢光顯微鏡觀測分離之效果，或以全反射螢光顯微技術(TIRF)來突破300 nm以下光學可觀測之粒徑極限。結合微粒子顯像測速儀量測系統與全反射螢光技術的應用，完整觀測奈米粒子與蛋白質鍵結之反應及其受介電泳力分離之過程。 預期本研究的成果將對於介電泳在奈米尺度生物分子的應用有直接的貢獻，尤其本研究將希望以光學觀測的方式來檢測生物分子反應後的狀況，可以達到非介入式之量測，對於不同型態之奈米生物粒子在非均勻電場下的行為直接觀測，有助於未來將此技術應用於生醫晶片中感測器之設計。 <br> Abstract: A novel sensing technique of bio-molecules is proposed based on the results developed in the previous project, Developments of Bio-sensor chip by detecting the Brownian motion of nano-particles. The antibodies linked on the nanobeads have rapid conjugation with the antigens in the buffer solution. The electric properties of the nanobeads may be changed after conjugating the antigens. For example, the surface charges or the polarization by an electric field will influence the behaviors of the nanobeads. In this project, the Dielectrophoresis (DEP) forces are employed to separate the nanobeads according to the conjugating statuses. The positive DEP or negative DEP forces depend on the properties of the nanobeads and the driving frequencies. The DEP separating technique is expected to easily identify the specific antigen in the buffer solution even the concentration of the antigen is low. Therefore, the high sensitive detection can be developed for the bio diagnostic system in the future. Based on the experiences of our previous studies, the biochips to separate the separations of normal and hardened red blood cells by DEP forces have been carried out. In order to separate the nano size beads, the NEMS fabrication technique is needed to manufacture the electrode array. In this study, the fluorescent nanobeads will be used to combine the proteins such as γ-globulin which exists for all human beings. The patterns of the electrode array will be designed when the electric properties are measured. The motions of the nanobeads in the electric field can be measured by an optical method by using micro Particle-Image-Velocimetry (μ-PIV) and Total Internal Reflection Fluorescence Microscopy (TIRFM). By experimental measurements and theoretical analysis with various parameters of the flow rates, frequency, and control voltages, the effects of particle-manipulating methods will be investigated in details. Results of this study can be used to better understand the flow phenomena to design Bio-chips and other MEMS applications by using dielectrphoresis force.奈米粒子介電泳粒子顯像測速儀量全反射螢光顯微鏡NanobeadsdielectrophoresisμPIVTIRF