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研究生: 陳燁萱
Yeh-hsuan Chen
論文名稱: 滲透壓對單層巨型微胞的影響
The Effect of Osmotic Pressure on Giant Unilamellar Vesicles
指導教授: 陳宣毅
Hsuan-yi Chen
口試委員:
學位類別: 碩士
Master
系所名稱: 理學院 - 生物物理研究所
Graduate Institute of Biophysics
畢業學年度: 99
語文別: 英文
論文頁數: 58
中文關鍵詞: 單層巨型微胞滲透壓形變
外文關鍵詞: Giant Unilamellar Vesicles, Osmotic Pressure, Shape deformation
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  • 本論文探討滲透壓對不飽和脂質(POPC)及不飽和脂質和飽和脂質以1:1比例(POPC:DPPC=1:1)組成的單層巨型微胞(GUVs,Giant Unilamellar Vesicles)的影響,GUVs以微胞電製法長出,將sorbitol加在GUVs外面,造成滲透壓,並且使用NBD-DOPE做為螢光分子,利用螢光顯微鏡觀察GUVs的形變。
    結果顯示不飽和脂質所組成的GUVs,加完濃度為0.225 M或0.3 M sorbitol後,一分鐘內GUVs直徑縮小幅度明顯並且可以觀察到許多小微胞在GUVs內部,之後達到穩定態。然而,我們選取直徑為35-45μm的GUVs,並其外面加0.3 M sorbitol,我們可以觀察許多小微胞在GUVs內部及外部;如果選取直徑大於70μm的GUVs, 一分鐘內GUVs縮小,之後會稍微膨脹,甚至於破裂。
    結果顯示不飽和脂質和飽和脂質以1:1比例(POPC:DPPC = 1:1)組成的GUVs擁有散亂或聚集的膠體區塊(gel phase domain),其行為模式有些不同。直徑小於40μm且擁有散亂的膠體區塊的GUVs,加sorbitol後,可以在GUVs裡面或是表面觀察到小微胞,且整個實驗過程GUVs維持類似球體的形狀; 直徑大於40μm且擁有散亂的膠體區塊的GUVs,加sorbitol後, 可以在GUVs裡面觀察到小微胞,GUVs的形狀變得不規則,有時GUVs甚至會破裂; 擁有聚集的膠體區塊的GUVs,加sorbitol後, 可以在GUVs裡面觀察到小微胞,並且整個實驗過程GUVs的形狀為不規則。


    In this work, we report experimental results on pure POPC and binary POPC/DPPC
    GUVs (Giant Unilamellar Vesicles) under osmotic pressure induced by sorbitol in the
    exterior solvent. GUVs were prepared by electroformation method. The evolution of
    GUVs was observed by
    uorescence microscopy using NBD-DOPE as the
    uorescent
    probe.
    The result for pure POPC GUVs is that GUVs usually shrink immediately and
    produce small vesicles inside themselves in the rst minute and then reach a steady
    state under osmotic pressure induced by 0:225 M or 0:3M sorbitol in the exterior
    solvent. However, under osmotic pressure induced by 0:3 M sorbitol in the exterior
    solvent, GUVs with initial diameter around 35
    􀀀
    45 m produce small vesicles both
    inside and outside the GUVs, and GUVs with diameter above70 m rst shrink then
    swell, eventually they burst.
    The results for binary POPC/DPPC GUVs with scattered or aggregated domains
    are quite di erent. GUVs with scattered domains and initial diameter below
    40 m produce small vesicles inside or on the outside surface of the GUVs and the
    shape of the GUVs are spheroidal under osmotic pressure, GUVs scattered domains
    and initial diameter above 40 m produce small vesicles inside and the shape of the
    GUVs become irregular, sometimes the GUVs may even burst. GUVs with aggregated
    domains produce small vesicles inside and the shape of the GUVs are irregular
    throughout the experiments.

    1 Introduction 1 1.1 The cell membrane . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 1.2 Shape deformations of giant unilamellar vesicles . . . . . . . . . . . . 4 1.3 Shape deformation coupled with phase behavior . . . . . . . . . . . . 6 1.4 Physics of vesicle deformation . . . . . . . . . . . . . . . . . . . . . . 6 2 Materials and methods 11 2.1 Materials . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11 2.2 Sample preparation . . . . . . . . . . . . . . . . . . . . . . . . . . . . 12 2.3 Giant unilamellar vesicles (GUVs) . . . . . . . . . . . . . . . . . . . . 13 2.4 Fluorescence microscopy . . . . . . . . . . . . . . . . . . . . . . . . . 14 2.4.1 The principle of uorescence microscopy . . . . . . . . . . . . 14 2.4.2 Fluorescence probe . . . . . . . . . . . . . . . . . . . . . . . . 14 2.4.3 The principle of uorescence . . . . . . . . . . . . . . . . . . 17 3 The e ect of osmotic pressure on pure POPC and POPC/DPPC binary GUVs 18 3.1 Homogeneous GUVs under osmotic pressure . . . . . . . . . . . . . . 19 3.1.1 Low sorbitol concentration (0:225 M) . . . . . . . . . . . . . 19 3.1.2 High sorbitol concentration (0:3 M) . . . . . . . . . . . . . . 22 3.2 Inhomogeneous GUVs under osmotic pressure . . . . . . . . . . . . . 27 3.2.1 GUVs with scattered domains . . . . . . . . . . . . . . . . . . 28 3.2.2 GUVs with aggregated domains . . . . . . . . . . . . . . . . . 33 4 Conclusions 38 5 Bibliography 42

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