研究動機：
胰臟癌 (Pancreatic cancer) 是一種臨床上難以早期診斷之癌症，且胰臟是人體消化系統中最重要的器官之一，癌細胞除了會造成胰臟中的脂肪酶、澱粉酶等滲漏而侵襲腹部鄰近的消化器官，也會快速轉移侵犯像是腸胃、肝和膽管等，導致常見的胰臟癌症狀有腹部疼痛、黃疸以及體重急降。因此，具有無警示性質、高轉移能力及化療抵抗力的胰臟癌，往往發現症狀時已經是晚期，五年存活率小於5%，然而目前唯一有效的治療方式只有早期診斷並手術切除。
研究目的：
因此，有其迫切及必要性來開發一種生物感測系統，可以快速準確地偵測早期胰臟癌生物因子。癌症抗原19-9 (CA19-9) 是一種由美國食品藥物管理署核准通過，最普遍應用於胰臟癌篩檢的癌指數，然而癌症抗原19-9在胰臟癌診斷上專一性 (82%) 及敏感性 (79%) 上仍顯不足，因此我們需要搭配另一種更具專一及敏感性的生物分子來開發我們的生物感測系統。黏液素1 (MUC1) 是一種在癌細胞膜表面上高度表現的蛋白，是目前最新研究上，針對早期胰臟癌檢測的新穎生物標記物，敏感性及專一性為82%及95%。
研究方法：
本研究中，發展了一種具備不同抗體 (anti-CA 19-9及anti-MUC1) 的磁性生物感測液體，可以快速且專一的捕捉到這兩種癌症抗原。固定了這兩種不同的抗體於奈米金桿上去作為第二層次的生物辨識及轉導元件，而且利用奈米金桿在強還原劑中能催化降解甲基橙試劑 (特定吸收光譜508 nm) 的特性，作為一衰減式比色光學生物感測系統。
研究結果：
在此設計中，當磁性感測液體固定的抗體捕捉到癌症抗原後，經磁石分離清洗，便加入96孔盤中，此時奈米金桿會透過三明治免疫的方式，結合到96孔盤底部的抗體上。最後，在反應完且清洗後的孔盤中，加入含有強還原劑－硼氫化鈉及甲基橙之溶液。當癌症抗原濃度越高時，磁性感測粒子就會捕捉到越多的抗原，而透過三明治免疫結合至孔盤底部的金桿就會隨之增多，因此，反應後催化降解甲基橙的效率也會提升。透過測定在不同濃度的癌症抗原標準品下，甲基橙吸收光的訊號強度下降趨勢，來分析出本系統的癌症抗原線性偵測範圍。

Motivation:
Pancreatic cancer (PC) represents a fatal tumor with a high mortality and poor prognosis. These outcomes can be attributed a lack of early diagnosis and inability to detect pre-cancerous pancreatic intraductal neoplastic (PanIN) lesions. Currently, no clinically reliable biosensing systems for early detection at an early stage of pancreatic cancer are available.
Purpose:
Thus, it’s urgent to develop a biosensing system which can fast and accurately diagnose PC. So far, the carbohydrate antigen (CA19-9) approved by the US Food and Drug Administration (FDA) is the most common biomarker used for several decades. However, it has limited value in diagnosis of early stages of PC. Mucin1 (MUC1) , a cell surface associated protein, is an emerging biomarker associated with early development of PC.
Methods:
Therefore, we develop a magnetic biosensing liquid modified with CA19-9 antibody and MUC1 antibody for the rapid and specific capturing of these two cancer antigens. Besides, we immobilize the anti-CA19-9 and anti-MUC1 onto the gold nanorods, which can catalyze the reduction of the methyl orange (Absorption at 508 nm) with the sodium bromide.
Results:
In this design, after the cancer antigens bind to the magnetic biosensing liquid, we add the liquid into the plate coated with antibodies to become sandwich immunosorbent conjugates. Finally, after washing the plate, we add the solution of methyl orange and sodium bromide into it.
Therefore, with increasing concentrations of cancer antigens, the more magnetic biosensing particles will bind onto the bottom of plate. And the efficiency of the catalytic degradation of methyl orange in a certain period of time will increase. In the end, we can use the downtrend of the absorption intensity under different concentrations of cancer antigens to analyze the linear detection range of our biosensing system.

目錄
審定書 i
誌謝 ii
中文摘要 iii
Abstract v
目錄 vii
圖目錄 ix
表目錄 xii
第1章 緒論 1
1-1 胰臟癌 1
1-1-1 概況 1
1-1-2 胰臟癌種類及成因 2
1-1-3 診斷 6
1-1-4 治療方式 10
1-2 生物感測器的介紹 11
1-2-1 不同類型之生物識別元件 12
1-2-2 不同類型之轉導元件 19
1-3 應用於胰臟癌抗原檢測之生物標誌及感測器 32
1-4 研究動機與目的 34
第2章 材料與方法 35
2-1 實驗藥品 35
2-2 儀器與設備 37
2-3 研究方法 38
2-4 奈米金桿膠體及改質不同分子的製備步驟 39
2-4-1 製備原理及過程 39
2-4-2 金桿表面牛血清蛋白的改質 40
2-4-3 以樹枝狀聚乙烯亞胺包覆之GNR@BSA製備 41
2-5 磁性奈米粒子製備及改質之步驟 41
2-5-1 磁性奈米粒子的製備 41
2-5-2 以牛血清蛋白包覆之磁性奈米粒子的製備 42
2-5-3 以樹枝狀聚乙烯亞胺包覆之Fe3O4@BSA的製備 43
2-6 牛血清蛋白改質及包覆bPEI材料催化甲基橙之光學測定實驗 43
2-7 磁性生物感測粒子及具抗體之塑膠孔盤的製備 44
2-7-1 磁性奈米粒子與半胱胺修飾之金桿複合物製備 44
2-7-2 半胱胺及樹枝狀聚乙烯亞胺金桿之催化能力比較 45
2-7-3 磁性生物感測粒子的抗體修飾 45
2-7-4 抗體固定於塑膠孔盤的製備流程 46
2-8 各種製備材料之鑑定分析及性質研究 47
2-8-1 吸收光譜分析 47
2-8-2 表面電位分析 47
2-8-3 穿透式電子顯微鏡 (TEM) 分析 48
2-8-4 傅立葉轉換紅外線光譜 (FTIR) 分析 48
2-8-5 X光高解析電子能譜儀 (XPS) 分析 48
2-8-6 超導量子干涉儀 (SQUID) 分析 48
2-8-7 基於化學發光 (Chemiluminescence) 的二抗免疫分析 49
2-9 CA19-9及MUC1癌症抗原的光學檢測 50
2-9-1 磁性生物感測粒子的催化能力測試 50
2-9-2 磁性生物感測粒子的穩定性測試 50
2-9-3 甲基橙之光學性質測試 50
2-9-4 單一抗原之光學檢測 50
第3章 結果與討論 52
3-1 奈米金桿膠體的製備及改質 52
3-1-1 不同體積硝酸銀對奈米金桿膠體溶液的影響 53
3-1-2 不同體積晶種液對奈米金桿膠體溶液的影響 55
3-1-3 金桿表面牛血清蛋白的改質 56
3-2 磁性奈米粒子的製備與改質 59
3-2-1 磁性奈米粒子的製備 60
3-2-2 磁性奈米粒子表面牛血清蛋白的包覆 61
3-3 催化甲基橙之光學測定實驗 65
3-3-1 牛血清蛋白改質材料之催化測試 66
3-3-2 以bPEI包覆之GNR@BSA及Fe3O4@BSA製備 68
3-3-3 包覆bPEI之材料催化測試 69
3-4 磁性生物感測粒子及具抗體之塑膠孔盤的製備 73
3-4-1 磁性奈米粒子與半胱胺修飾之金桿複合物製備 73
3-4-2 磁性生物感測粒子的抗體修飾 77
3-4-3 抗體固定於塑膠孔盤的製備 80
3-5 CA19-9及MUC1癌症抗原的光學檢測 83
3-5-1 磁性生物感測粒子的催化能力測試 83
3-5-2 磁性生物感測粒子的穩定性測試 87
3-5-3 單一抗原之光學檢測 88
第4章 結論 91
參考文獻 92

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