臺灣博碩士論文加值系統

鉤端螺旋體病是人畜共通疾病，是屬於鉤端螺旋體屬的螺旋體菌。此疾病可由鉤端螺旋體污染的動物排泄物、水、土壤經由皮膚、眼、口或鼻感染。此疾病的臨床特徵從輕微至嚴重的發燒，有時甚至是致命的疾病，如腎衰竭，黃疸，出血（特別是影響肺）和血管塌陷。而世界衛生組織認為鉤端螺旋體感染患者的死亡率可能高達5％至25％。目前的黃金診斷標準是微量凝集試驗(MAT)、酶聯免疫測定法（ELISA）或聚合脢連鎖反應(PCR)則需貴重精密檢測儀器且需專業技術人員。本研究的目標是要達成快速、簡易並有效率的方法，檢測標的細菌-鉤端螺旋體菌。我們設計一個細菌捕捉器，以聚二甲基矽氧烷(PDMS)製作一個漏斗形狀的腔室，貼於蓋玻片上，並在腔室底部鍍上鉤端螺旋體專用陽性血清，再利用水平離心原理，於在短時間內快速捕獲低濃度的鉤端螺旋體。本研究之實驗結果預期將有助於鉤端螺旋體之診斷、觀察和研究。

Leptospirosis is a worldwide zoonotic disease caused by pathogenic spirochetes of the genus Leptospira. This disease is infected by animal excrement, water, soil containing Leptospira coming into contact with breaks in the skin, eyes, mouth, or nose. Clinical symptoms range from a self-resolving acute undifferentiated febrile illness to severe, sometimes fatal disease with renal failure, jaundice, hemorrhage (particularly affecting the lungs), and vascular collapse. WHO believes the leptospirosis death rate may be between 5% to 25% of infected patients. The gold standard method of Leptospirosis diagnosis is the micro-agglutination test (MAT) that is time consuming. New molecular diagnostic methods, including polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA) have high sensitivity and provide fast diagnosis. However, they require high-tech equipments and experts. Except the molecular diagnostic methods, one possible method is to directly capture Leptospira from samples with a slide which is coated specific antibodies. However, it is difficult to implement because the Leptospire is very active. The goal of this study is to develop a rapid and simple detection method for capturing Leptospira using a special designed chamber assembled with a funnel-shaped chamber made with Polydimethylsiloxane (PDMS) and a cover slide. The positive serum was coated on the bottom of the chamber. Using a horizontal centrifugal, the bacteria capture can be reached in a short time. The operation of the antibody coating is time consuming, and it needs a considerable quantity of materials. This study is expected to contribute to the diagnosis and research of the Leptospirosis.

中文摘要 I
Abstract II
目錄 III
圖目錄 VI
表目錄 VIII
第一章 緒論 1
1.1研究動機及背景 1
1.2研究目的 2
1.3論文架構 3
第二章 文獻回顧 5
2.1鉤端螺旋體病的發現 5
2.1.1鉤端螺旋體種類 6
2.1.2鉤端螺旋體病傳染、感染途徑與症狀 8
2.2鉤端螺旋體病診斷方法 10
2.2.1鉤端螺旋體分離與鑑定 10
2.2.2鉤端螺旋體抗體檢測-顯微凝集法 13
2.2.3鉤端螺旋體抗體檢測-酵素免疫分析法 16
2.2.4鉤端螺旋體抗體檢測-螢光定量PCR 19
2.2.5鉤端螺旋體抗體檢測-等溫擴增定量檢測 20
2.2.6鉤端螺旋體檢測結論及比較 21
2.3生物感測晶片 24
2.3.1微陣列法 24
2.3.2基因晶片法 25
2.3.3蛋白質/抗體晶片 26
2.3.4組織晶片 27
2.3.5實驗室晶片 28
2.4光學顯微鏡 29
2.4.1相位差顯微鏡 29
2.4.2暗視野顯微鏡 30
第三章 材料與方法 31
3.1實驗設備 31
3.1.1桌上型離心機 31
3.1.2倒立式顯微鏡 33
3.1.3 3D Printer 33
3.2 樣品介紹及備製 34
3.2.1細菌捕捉器材料 34
3.2.2抗體鍍膜 35
3.2.3鉤端螺旋體及抗體 35
3.3 實驗方法 37
3.3.1實驗所需材料 37
3.3.2實驗流程 38
第四章 結果與討論 40
4.1 PDMS細菌捕捉器之研製 40
4.2實驗過程之鉤端螺旋體觀察 43
4.3離心參數之測試 47
4.4鍍膜實驗 51
第五章 結論 56
參考文獻 58

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