研究生: |
林奕儒 I-Ju Lin |
---|---|
論文名稱: |
鮑氏不動桿菌之紫膜生物光電感測晶片的開發 Development of purple membrane-based photoelectric chips for Acinetobacter baumannii detection |
指導教授: |
陳秀美
Hsiu-Mei Chen |
口試委員: |
吳雪霞
Hsueh-Hsia Wu 葉旻鑫 Min-Hsin Yeh |
學位類別: |
碩士 Master |
系所名稱: |
工程學院 - 化學工程系 Department of Chemical Engineering |
論文出版年: | 2021 |
畢業學年度: | 109 |
語文別: | 中文 |
論文頁數: | 138 |
中文關鍵詞: | 鮑氏不動桿菌 、紫膜 、生物傳感器 、生物光電晶片 |
外文關鍵詞: | Acinetobacter baumannii, purple membrane, biosensor, photoelectric chips |
相關次數: | 點閱:212 下載:0 |
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鮑氏不動桿菌 (Acinetobacter baumannii)為造成嚴重院內感染之病原體,其能引起肺炎、腦膜炎,甚至是高死亡率的菌血症,因此快速且靈敏的辨別出A. baumannii蔚為重要,唯有即時鑑定病原體才能增加施藥準確率,且避免用藥錯誤產生抗藥性並能有效降低死亡率。細菌視紫質 (bacteriorhodopsin, BR)存在於古生嗜鹽菌 (Halobacterium salinarum)的紫色細胞膜 (purple membrane, PM)中,其為受光激發會產生跨膜質子梯度差的光驅動質子泵,將此特性透過外接電路即可量測到光電流訊號。本論文即運用以PM為光電訊號轉換器之原理所製備的PM生物感測晶片,以針對A. baumannii具有高度專一性的核酸適體為辨識因子,開發A. baumannii aptamer-PM複合生物光電感測晶片,用以檢測A. baumannii。首先對晶片製程中核酸適體架橋的種類與長度、檢測緩衝液中Na+濃度進行探討,尋求對檢測效果最為顯著之製備條件。如此以最佳製程製備的A. baumannii aptamer-PM複合生物光電感測晶片可檢測A. baumannii最低濃度至1E-1 CFU/mL,接續測試其對台灣常見七種菌血症菌株之分辨能力,結果顯示本論文開發的A. baumannii aptamer-PM複合生物光電感測晶片對A. baumannii具有顯著區別能力 (p<0.01,具有顯著差異性)。最後再利用結合辨識因子的奈米金進行訊號增強的探討,發現如此製程對A. baumannii之檢測限能降低至濃度1E-2 CFU/mL。
Acinetobacter baumannii is a pathogen causing severe nosocomial infection, resulting in meningitis, pneumonia, or even lethal bacteremia. Therefore, the rapid and sensitive identification of A. baumannii is important for precision antibiotics to avoid the development of antibiotic resistance and to reduce the mortality rate. Bacteriorhodopsin (BR) exists in the purple membrane (PM) of Halobacterium salinarum and acts as a light-driven proton pump, generating a cross-membrane proton gradient which drives photocurrents through an external circuit. This thesis employs a previously developed sensor with PM as a photoelectrical signal transducer to further devise an aptamer-PM complex biosensor for A. baumannii detection. The biosensor contains specific aptamers against A. baumannii as the recognition element. For optimal biosensor development and detection, different aptamers were investigated, as well as the lengths of their conjugation linkers with PM and the Na+ concentration in the detection buffer. The best developed A. baumannii aptamer-PM complex biosensor sensitively detected A. baumannii down to 1E-1 CFU/mL and significantly discriminated A. baumannii from 6 other common bacteremia-causing strains found in Taiwan (p < 0.01). Finally, to further enhance the detection sensitivity, gold nanoparticles conjugated with the optimal recognition aptamer were used to label the microbes captured on the A. baumannii aptamer-PM complex sensing chip, resulting a detection limit of 1E-2 CFU/mL.
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