原子力顯微鏡（Atomic force microscope ; AFM）對表面形貌具有高解析度的成像能力，也能探測奈米尺度的力學性質，因此成為表面科學的標準研究工具之一。本文採用兩種不同分化程度的人類肝癌細胞，即SK-HEP-1 （分化程度低、癌化程度高）、 Hep G2 （分化程度高、癌化程度低）的肝癌細胞，將其固定於玻璃基板上，以AFM 探究細胞表面的形貌，並記錄懸臂偏折與應力大小之力曲線圖（force curve），可以有效表現出作用力與壓痕深度的關係。同時，考慮細胞表面的非均質性，故將細胞表面不同局部位置取得的力曲線作為單一影像點（pixel），以pixel by pixel 的影像點陣列型式，組合成表面力學特性的分佈圖（force map），以比較兩種細胞表面不同局部位置在彈性（elasticity）、黏著力（adhesion force）等奈米力學性質上的相對差異；最後，藉由Hertzian model 的計算方式，計算兩種細胞之楊氏係數（Young's modulus）的大小。結果顯示，兩種細胞在表面形貌、高度、細胞的遷移性、細胞與聚集生長的程度、細胞表面的粗糙度、細胞表面彈性的性質、細胞表面的黏著力上均有所差異。SK-HEP-1 的細胞
邊緣皺褶的間隔距離較寬、細胞遷移性較佳、細胞表面呈現軟硬程度較為均質的彈性特性，可以假設其在細胞膜下具有密集的網狀分枝 ( Branched networks ) 結構；而 Hep G2 的細胞邊緣皺褶的間隔距離較緊密、細胞遷移性較差、細胞表面呈現軟硬程度較不均質的彈性特性，可以假設其在細胞膜下具有分散的肌動蛋白絲與網狀交聯 ( Cross-linked networks ) 兩種細胞骨架結構。綜合以上結果，可以推測樣品表面的彈性性質會受到細胞膜下肌動蛋白骨架的影響，能假設細胞中細胞骨架的結構與其分布的情形，此假設符合實驗結果的驗證。

Atomic force microscopy (AFM) can be used to acquire high-resolution topographical images of surfaces, but has the additional capability of detecting the local nanometer scale mechanical properties. For these reasons, it becomes a standard research tool in the surface science recently. In this paper, we used AFM to measure the several properties of two different human hepatocellular
carcinoma cell lines, Hep G2 ( known as well differentiated and more highly carcinomatous hepatoma cell lines ) and SK-HEP-1 ( known as poorly differentiated and more lightly carcinomatous hepatoma cell lines ) cells fixed on the glass substrate, which including the surface morphology and the relationship between the cantilever deflections and loading forces ( force curve ). Considered the heterogeneous characteristics of the cell surface, the
preferred experimental method is to make pixel-by-pixel force curves in a designated area ( force map ) , both adhesion forces and elasticity associated with different locations on the cell surfaces will be obtained. Finally, we use Hertzian model to calculate Young's modulus of Hep G2 and SK-HEP-1 respectively. Based on these results, we can understand the surface properties of two human hepatocarcinoma cell lines with different differentiated stage. The results showed the difference of the morphology, height, cell migration, degree of cell aggregation, roughness, elasticity, adhesive force of two cells. SK-HEP-1 cell has the wide distance of the folds, better cell migration, homogeneous properties of elasticity. It can be assumed that the SK-HEP-1 cells have a dense network structure of actin filaments under the cell membrane like branches (branched networks); Hep G2 cell has the narrow distance of the folds, poor cell migration, heterogeneous properties of elasticity. It can be assumed that the Hep G2 cells have the individual actin
filaments and cross-linked network structure of actin filaments under the cell membrane. The above results can be speculated that the elastic properties of the membrane surface will be influenced of actin filaments.

中文摘要………………………………………………………… i
英文摘要………………………………………………………… iii
目錄……………………………………………………………… v
圖目錄…………………………………………………………… vii
表目錄…………………………………………………………… xi
第壹章 緒論 …………………………………………………… 1
1-1 研究背景…………………………………………………… 1
1-2 文獻回顧…………………………………………………… 3
1-2-1 生物樣品顯微觀測技術…….………..…………….. 3
1-2-2 原子力顯微鏡觀測生物樣品.………..…………….. 4
1-2-3 細胞骨架.………..……………..…………………… 6
1-3 研究動機..………………………………………………… 10
第貳章 實驗儀器與實驗方法……………………………… 12
2-1 實驗儀器..………………………………………………… 12
2-1-1 光學顯微鏡………………….………..…………… 12
2-1-2 原子力顯微鏡………………….………..………… 13
2-2 生物樣品製備與原子力顯微鏡的觀察…………………… 20
2-2-1 人類肝癌細胞株…………….………..…………… 20
vi
2-2-2 細胞培養…………………….………..…………… 21
2-2-3 原子力顯微鏡觀察細胞樣品.………..…………… 21
2-3 實驗數據的分析方法……………………………………… 23
2-3-1 Hertzian model.………..…………………………… 23
2-3-2 原子力顯微鏡的力曲線圖.………………………… 25
2-3-3 細胞表面彈性的分析方式………………………... 27
2-3-4 細胞表面黏著力的分析方式……………………… 29
第參章 結果與討論…………………………………… 32
3-1 光學顯微鏡……….………………..……………… 32
3-2 AFM 測量細胞表面形貌……………..……………... 33
3-3 AFM 測量細胞表面力學性質……..………………... 38
3-3-1 細胞表面彈性性質……..…………………………... 40
3-3-2 細胞表面黏著力……..…………………………....... 44
3-4 實驗結果與推論……..……………………………….. 49
3-4-1 實驗結果……..……………………………………... 49
3-4-2 細胞結構對表面性質的影響……………………..... 51
第肆章 結論………………………………………..……… 57

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