膠原蛋白為生物體的結構型蛋白質，例如：肌肉組織、骨骼和韌帶等，在動物體中所扮演的角色為連接細胞與組織。而「溫度」這個與我們生活習習相關的天然環境變數，則是深深地影響到膠原蛋白的功能與型態。在生物學的研究中，溫度引起之膠原蛋白結構改變是一重要主題。傳統上，研究的方式大多係使用差分掃瞄熱量儀DSC與非線性光學偵測型變(conformational change)的產生。DSC可以量測出膠原蛋白的相變點、反應熱與熱穩定性等熱力學性質，但缺點則是無法產生局部化之資訊。使用非線性倍頻光學的方式，則可以藉由倍頻光的強度推測得膠原蛋白結構的變化，並且可以產生倍頻影像，進行局部變化的分析。在之前的研究成果中發現到，環境中的水分子濃度關係到膠原蛋白結構對溫度變化的穩定性。相對地，本研究使用一真空變溫平台以控制環境，藉以隔絕環境中的水分子、氧氣與雜質。配合以非線性掃瞄式倍頻光學顯微鏡的使用，便可以即時變溫的方式觀察真空中膠原蛋白的變化，並藉由倍頻顯微影像的強度，推測出膠原蛋白結構的改變。

Collagen is an important structural protein in living organisms and plays an indispensable role in connecting cells and tissues, such as in musculature, bone, and ligament. The stability and conformation of collagen are, however, strongly influenced by ambient temperature and constitutes an interesting subject of study. Thermally induced conformation change of collagen has been investigated by techniques such as differential scanning calorimetry (DSC) and second harmonic generation. DSC is a powerful method in uncovered important thermal dynamics properties including phase change, enthalpy, and thermal stability of the collagen. However, due to its collective nature, no localized information can be found. For comparison, second harmonic generation, which reflects structural symmetry, can be combined with laser scanning microscopy to investigate localized variation. It has been shown in previous studies that the thermal stability of collagen is strongly influenced by the water content within collagen. For comparison, we are investigating the conformational change of collagen under a vacuum stat with second harmonic microscopy so as to isolate environmental effects, particularly those from water and oxygen. In this way, we have found the conformational change of collagen takes place at a much higher temperature and activation energy. Additionally, the high spatial resolution achieved also allows many further possibilities.

第一章　緒論．．．．．．．．．．．．．．．．．．．．．．．01
第二章　二倍頻顯微術理論與膠原蛋白結構
2.1非線性光學倍頻簡介．．．．．．．．．．．．．．．．04
2.2蛋白質與膠原蛋白之結構．．．．．．．．．．．．．．06
第三章　研究方法
3.1樣本選取．．．．．．．．．．．．．．．．．．．．．13
3.2研究架構．．．．．．．．．．．．．．．．．．．．．13
3.3實驗步驟．．．．．．．．．．．．．．．．．．．．．15
3.4資料處理．．．．．．．．．．．．．．．．．．．．．16
第四章　結果與討論
4.1倍頻訊號與溫度關係．．．．．．．．．．．．．．．．18
4.2不同溫度下，倍頻訊號隨時間的衰減．．．．．．．．．19
第五章　結論．．．．．．．．．．．．．．．．．．．．．．．23
參考文獻．．．．．．．．．．．．．．．．．．．．．．．．．24
附錄A　20種氨基酸化學結構．．．．．．．．．．．．．．．．27
附錄B　雷射光源系統．．．．．．．．．．．．．．．．．．．28
附錄C　電子偵測系統．．．．．．．．．．．．．．．．．．．31
附錄D　真空變溫系統．．．．．．．．．．．．．．．．．．．34
附錄E　光學元件．．．．．．．．．．．．．．．．．．．．．35
附錄F　掃描式共焦顯微系統．．．．．．．．．．．．．．．．37

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