人類白蛋白為人體循環系統中最豐富的蛋白質，由肝臟所製造，分子量為67千道爾吞，由 585個胺基酸所組成。普遍用來作為檢測尿液的生物標記物，檢測是否患有高血壓或糖尿病所造成的慢性腎臟病，以及病人的腎功能是否逐漸受損的一個指標。
  在我們的研究中，我們使用液晶在偏折光下的特殊光學訊號作為讀出系統來發展液晶型免疫分析系統，用以檢測人類白蛋白在水溶液中的含量。這種分析方法的檢測機制是當人類白蛋白與玻璃表面上的人類白蛋白抗體進行特異性結合時造成玻璃表面的蛋白質密度不同，而導致液晶排列方向的不同而在偏折光下有暗到亮的圖像轉換。我們的結果顯示這種液晶型免疫分析系統可以檢測人類白蛋白的濃度為1 μg/mL。

Human serum albumin (HSA) is the most abundant protein in human circulatory system. It is produced in the liver. It molecular weight is 67 kDa, consists of 585 amino acids. It is commonly used as a biomarker in urine samples for detecting chronic kidney disease (CKD) caused by high blood pressure or diabetes, a condition in which a patient lose their kidney function gradually. 
    In our research, we used a thin layer of liquid crystals (LCs) is used as a readout system for developing an immunoassay that reports the presence of HSA in the aqueous solution with optical signals. The detection mechanism of this assay is based on the variation of surface density of protein upon the specific binding of HSA on anti-HSA immobilized surface, which lead a dark-to-bright transition of LC images under cross polarizers. Our results show that the LC-based assay can detect HSA at concentration of 1 μg/mL.

目錄
中文摘要.....................................................................................................I
英文摘要...................................................................................................II
目錄..........................................................................................................III
圖表目錄..................................................................................................IV
第一章 緒論..............................................................................................1
 1-1 血清白蛋白.......................................................................................2
 1-2 以血清白蛋白作為慢性腎臟病指標...............................................3
 1-3 血清白蛋白的分析方法...................................................................5
  1-3-1 免疫分析法.................................................................................5
  1-3-2 尿液試紙...................................................................................12
 1-4 液晶型分析方法.............................................................................13
  1-4-1 液晶...........................................................................................13
  1-4-2 液晶型分析方法的檢測機制...................................................16
  1-4-3 液晶型分析方法的介紹...........................................................18
   1-4-3-1 液晶-固體介面.....................................................................18
   1-4-3-2 液晶-水溶液介面.................................................................20
   1-4-3-3 液晶液滴..............................................................................22
  1-4-4 液晶型分析方法的優點...........................................................23
 1-5 液晶型免疫分析系統.....................................................................24
 1-6 總結.................................................................................................29
第二章 實驗方法與實驗儀器................................................................31
 2-1 實驗藥品與材料.............................................................................31
 2-2 實驗儀器.........................................................................................32
 2-3 實驗目的與實驗方法.....................................................................33
  2-3-1 實驗目的...................................................................................33
  2-3-2 實驗方法...................................................................................33
   2-3-2-1 製備不同官能基修飾的載玻片..........................................33
    2-3-2-1-1 製備DMOAP修飾的載玻片.........................................33
    2-3-2-1-2 製備醛基官能基修飾的載玻片.....................................34
    2-3-2-1-3 製備胺基官能基修飾的載玻片.....................................34
    2-3-2-1-4 製備鏈黴抗生物素蛋白修飾的載玻片.........................35
  2-3-2-2 製備蛋白質溶液....................................................................35
   2-3-2-2-1 製備血清白蛋白溶液.......................................................35
   2-3-2-2-2 製備抗血清白蛋白溶液...................................................36
   2-3-2-2-3 製備氧化抗血清白蛋白溶液...........................................36
  2-3-2-3 製備抗血清白蛋白晶片........................................................37
   2-3-2-3-1 抗血清白蛋白晶片...........................................................37
   2-3-2-3-2 氧化抗血清白蛋白晶片...................................................37
   2-3-2-3-3 Affibody 晶片................................................................38
  2-3-2-4 液晶型免疫分析....................................................................38
   2-3-2-4-1 抗體與抗原結合力測試...................................................39
   2-3-2-4-2 檢測血清白蛋白...............................................................40
	2-3-2-4-3 氯化鎂對於檢測血清白蛋白的影響...............................40
   2-3-2-4-4 聚乙烯亞胺對於檢測血清白蛋白的影響.......................41
   2-3-2-4-5 二級抗體對於檢測血清白蛋白的影響...........................41
  2-3-2-5 製備液晶元件........................................................................42
  2-3-2-6 液晶元件成像........................................................................43
  2-3-2-7 FT-IR 測定載玻片表面官能基..............................................43
第三章 結果與討論................................................................................44
 3-1 不同官能基修飾載玻片的液晶光學訊號圖像.............................44
  3-1-1 DMOAP修飾的載玻片..............................................................44
  3-1-2 醛基修飾的載玻片...................................................................48
  3-1-3 胺基修飾的載玻片...................................................................49
 3-2 界面活性劑Tween 20對於檢測血清白蛋白的影響....................51
  3-2-1 Tween 20的最佳濃度條件........................................................51
 3-3 抗體與抗原結合力測試.................................................................53
  3-3-1 血清白蛋白與抗血清白蛋白結合力測試...............................53
  3-3-2 血清白蛋白與氧化抗血清白蛋白結合力測試.......................55
 3-4 固定抗血清白蛋白.........................................................................56
 3-5 血清白蛋白檢測.............................................................................58
 3-6 氯化鎂對於檢測血清白蛋白的影響.............................................60
  3-6-1 氯化鎂的最佳濃度條件...........................................................60
  3-6-2 血清白蛋白加入氯化鎂後的結合力測試...............................62
  3-6-3 氯化鎂對於檢測血清白蛋白的影響.......................................64
 3-7 抗血清白蛋白晶片的專一性免疫結合測試.................................65
 3-8 聚乙烯亞胺對於檢測血清白蛋白的影響.....................................67
 3-9 二級抗體對於檢測血清白蛋白的影響.........................................68
  3-9-1 單株二級抗血清白蛋白的結合力測試...................................69
  3-9-2 多株二級抗血清白蛋白的結合力測試...................................70
  3-9-3 單株二級抗血清白蛋白對於檢測血清白蛋白的影響...........71
  3-9-4 多株二級抗血清白蛋白對於檢測血清白蛋白的影響...........73
  3-9-5 含生物素二級抗血清白蛋白對於檢測血清白蛋白的影響...74
 3-10利用AffibodyR 晶片檢測血清白蛋白........................................76
第四章 結論............................................................................................78
第五章 參考資料....................................................................................82
圖表目錄
圖1-1 免疫分析法基本原理....................................................................5
圖1-2 免疫分析法類型............................................................................6
圖1-3 典型尿液試紙..............................................................................13
圖1-4 液晶相圖......................................................................................14
圖1-5 液晶的特殊光學訊號..................................................................15
圖1-6 4-戊基-4'-氰基聯苯的結構...........................................................16
圖1-7 液晶型分析方法檢測機制..........................................................17
圖1-8 顯微鏡中交叉偏光板的原理示意圖..........................................18
圖1-9 液晶−固體介面............................................................................20
圖1-10 DMOAP結構..............................................................................20
圖1-11 液晶−水溶液介面......................................................................21
圖1-12 液晶液滴....................................................................................22
圖1-13 液晶光學訊號............................................................................26
圖1-14 液晶元件裝置圖........................................................................28
圖1-15 蛋白質吸附於表面後造成液晶分子排列方向改變................28
圖1-16 血清白蛋白檢測晶片................................................................30
圖2-1 氧化抗血清白蛋白示意圖..........................................................36
圖2-2 檢測流程......................................................................................39
圖2-3 製備液晶元件流程......................................................................42
圖3-1 DMOAP修飾的載玻片液晶光學訊號圖與表面修飾示意圖....45
圖3-2 FT-IR測定載玻片表面DMOAP結果圖......................................47
圖3-3 醛基修飾的載玻片液晶光學訊號圖與表面修飾示意圖..........49
圖3-4 胺基修飾的載玻片液晶光學訊號圖與表面修飾示意圖..........50
圖3-5 界面活性劑Tween 20的最佳化濃度條件與氧化抗血清白蛋白晶片示意圖.................................................................................52
圖3-6 結合力測試流程圖......................................................................53
圖3-7 結合力測試結果與實驗示意圖..................................................54
圖3-8 氧化抗血清白蛋白結合力與活性測試結果與實驗示意圖......56
圖3-9 固定抗血清白蛋白的結果與製備示意圖..................................58
圖3-10 檢測不同濃度血清白蛋白的結果與檢測示意圖....................60
圖3-11 氯化鎂最佳濃度結果與實驗示意圖........................................62
圖3-12 血清白蛋白溶液加入氯化鎂結合力測試與實驗示意圖........63
圖3-13 緩衝溶液加入氯化鎂的檢測結果與實驗示意圖....................65
圖3-14 特異性免疫結合測試結果與實驗示意圖................................66
圖3-15 緩衝溶液加入聚乙烯亞胺的檢測結果與實驗示意圖............68
圖3-16 單株二級抗血清白蛋白的結合力測試與實驗示意圖............70
圖3-17 多株二級抗血清白蛋白的結合力測試與實驗示意圖............71
圖3-18 加入單株二級抗血清白蛋白檢測結果與實驗示意圖............72
圖3-19 加入多株二級抗血清白蛋白的檢測結果與實驗示意圖........74
圖3-20 加入含生物素二級抗血清白蛋白的檢測結果與實驗示意....75
圖3-21 AffibodyR 晶片檢測結果與實驗示意圖...................................77
表1-1 慢性腎臟病之分期........................................................................4
表2-1 實驗藥品與材料..........................................................................31
表4-1 血清白蛋白檢測法比較..............................................................80

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