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標題: | 原子力顯微術於流感病毒之表面奈米結構與奈米力學之研究 Investigation of Surface Nanostructure and Nanomechanics of Influenza Virus using Atomic Force Microscopy |
作者: | Chih-Hao Liu 劉志豪 |
指導教授: | 林世明(Shiming Lin) |
關鍵字: | 原子力顯微術,麻杏石甘湯,表面粗操度,定位,解離力,流感病毒, Atomic force microscopy (AFM),MXSGT,Surface roughness,Localization,Unbinding force,Influenza virus, |
出版年 : | 2012 |
學位: | 博士 |
摘要: | 原子力顯微鏡(AFM)是奈米生物科技研究上的一個重要工具,其所得到的影像具有奈米等級的空間解析度,同時於力學應用上,可達到皮米牛頓等級的靈敏度和奈米等級之定位精度。
本研究利用AFM作為檢測平台,首先由流感病毒顆粒的表面結構開始探討,並利用不同濃度的中草藥麻杏石甘湯(MXSGT)對於病毒進行處理,藉由AFM所獲得的高解析度影像,來分析處理前後的流感病毒顆粒表面形貌並得到其表面粗糙度參數。本研究成功的利用病毒顆粒表面之3D影像、相位圖以及四組表面粗糙度參數,達到分析控制組與不同濃度麻杏石甘湯處理後的奈米等級差異。 接下來,本研究利用高專一性的抗體配合化學修飾共價鍵結於AFM之探針形成抗體探針,接著整合了前述由AFM影像所獲得之病毒顆粒幾何參數,成功地建立一套三階段的病毒顆粒的定位技術。藉由這項技術,除了可量測單顆病毒表面抗原及抗體探針間的力學參數,並可同時繪製單顆病毒顆粒的表面抗原分佈,本研究也透過實驗及統計分析驗證了此定位技術的重複性。藉由這樣的技術,可針對單顆病毒顆粒的力學性質提出一系列量化數據,由於這項技術不需要外加硬體配件及特定的系統配置,可以很容易地適用於大多數的AFM系統,也因此增進AFM研究特定病毒表面抗原之能力。 因此,透過本研究所累積的成果與開發的相關技術,可提供未來一個新的面向,以病毒顆粒為單位來研究經藥物處理後之病毒形貌,以及抗體與病毒表面抗原力學的量化差異。 Atomic force microscopy (AFM) is a vital instrument in nanobiotechnology that not only enables imaging with nanometer spatial resolution but also can determine the interaction force of a single biomolecule with pico-Newton force sensitivity and nanometer positional accuracy using functionalized probes. In this thesis, firstly, AFM was used to visualize and quantify the effects of the Chinese herbal medicine Ma-xing-shi-gan-tang (MXSGT) on the surface topography of influenza virus particle, accompanied by high-resolution AFM images of cultured influenza virus and virus treated with various concentration of MXSGT. We then used the surface roughness parameters extracted from AFM images to quantitatively access the nanoscale differences on the outer surface of virus particles and statistically validate the differences between the control and treatment groups. Next, we developed a method that enables AFM to simultaneously measure specific unbinding force and map the viral glycoprotein at the single virus particle level. Based on the purposed method and performed analysis, several findings can be derived from the results. The mean unbinding force of a single virus particle can be quantified, and no significant difference exists in this value among virus particles. Furthermore, the repeatability of the proposed method is demonstrated. The force mapping images reveal that the distributions of surface viral antigens recognized by antibody probe were dispersed on the whole surface of individual virus particles under the proposed method and experimental criteria. This approach can be easily applied to most AFM systems without specific components or configurations. These results help understand the force-based analysis at the single virus particle level, and therefore, can reinforce the capability of AFM to investigate a specific type of viral surface protein and its distributions. In summary, the obtained results and developed method in this research offers new prospects for analyzing the treatment effects on nanoscale viral topography and the force interactions between antibody and viral surface antigen at a single virus particle level. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/66324 |
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顯示於系所單位: | 應用力學研究所 |
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