https://scholars.lib.ntu.edu.tw/handle/123456789/179095
標題: | 比率計數螢光法偵測pH值與追蹤癌細胞內單一粒子 Ratiometric Fluorescence pH Sensing and Single-particle Tracking in Cancer Cell |
作者: | 朱佳音 Chu, Jia-Yin 牟中原指導 |
關鍵字: | 奈米矽材;比率計數螢光法;細胞內酸鹼值;單分子追蹤;silica nanoparticles;ratiometric fluorescence;intracellular pH value;single-particle tracking | 公開日期: | 2012 | 摘要: | 細胞內的pH值於細胞的各個功能上扮演著重要的腳色。因為細胞內多數的蛋白質的性質會因為氫離子濃度的不同而有所改變;而且有部分的反應機制會被細胞內的pH值所調控著。因此,使用空間上高解析度的技術作為觀測pH值的手段,可以幫助我們更加了解細胞內部生理以及病理的過程。 我們合成大小為100 nm具有中孔性質的奈米矽材,在矽材裡添加兩種螢光染料,其中一種能夠對環境中的pH值做感測, FITC;另外一種則無法對pH值做感測, RITC。藉由將FITC的放光強度除以RITC放光強度所得的比值來估計奈米矽材所在環境的pH值。此外,提高螢光染料的附載量來增加材料的光穩定性。最後在奈米矽材表面修飾上帶有不同電性的矽烷;即可獲得在其表面帶有正電以及負電的奈米矽材。 接著,我們利用單分子追蹤系統 (single-particle tracking system) 來觀察帶有兩種不同表面電荷的奈米矽材,被癌細胞所吞噬過程或在細胞膜上移動的軌跡。此外,因為我們奈米矽材能做pH值的感測,所以除了獲得這些軌跡之外,我們還可以同步地得到這些軌跡裡每一點的環境中pH值。 The intracellular pH is an important modulator of cell functions. Since the activities of most proteins are affected by very small changes of the proton concentration, there are a number of cellular mechanisms that finely regulate the intracellular pH values (pHi). Therefore, monitoring pHi with high spatial resolution could help us to elucidate many physiological or pathogenic processes taking place within cells. We synthesized 100 nm mesoporous silica nanoparticles with a pH-sensitive dye, FITC and a pH-insensitive dye, RITC. This nanoparticle was employed to estimate the pHi by the fluorescence intensity ratio of FITC to RITC. We conjugated high amount of dyes to improve the photostability of the dyed nanoparticles. Besides, we modified the surface by post-modification to form two different nanoparticles, positively charged TA-hMSN and negatively charged THPMP-hMSN. The nanoparticles were characterized with various physical methods. Then, we used the single-particle tracking (SPT) system to examine the interaction between the nanoparticles and HeLa cells. We obtained the trajectories of nanoparticles during the uptake process and were able to estimate the fluorescence ratio in the local environment simultaneously. |
URI: | http://ntur.lib.ntu.edu.tw//handle/246246/257229 |
顯示於: | 化學系 |
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ntu-101-R99223112-1.pdf | 23.32 kB | Adobe PDF | 檢視/開啟 |
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