古生嗜鹽菌Halobacterium salinarum 具有一種名為紫膜（purple membrane, PM）的細胞膜，內含細菌視紫質（bacteriorhodopsin, BR）光敏感蛋白，其受光激發後會產生膜內外質子濃度差，可用以生產光電流。利用微生物會散射光與PM光電流強度與入射光強度呈線性關係之事實，本論文開發具有高靈敏度可定量檢測Streptococcus mutans牙斑菌PM生物感測晶片。本研究分別以protein A定向化之S. mutans抗體與對S. mutans具專一性且以化學鍵結固定化之核酸適體為辨識分子固定化於PM晶片上，製作出兩種PM複合感測晶片，並比較在靜態以及微流道內對S. mutans的檢測結果。除了觀察晶片光電流訊號變化外，並同步以螢光顯微鏡確認菌體有被捕捉在晶片上。於靜態檢測時，抗體-PM複合晶片對S. mutans的檢測靈敏度可低至10 CFU/mL；核酸適體-PM複合晶片的檢測靈敏度則更可至1 CFU/mL，兩者的檢測範圍均至少可達106 CFU/mL。比較靜態檢測與微流道內即時檢測，於高菌體濃度時，兩種檢測方式有同樣效果；然而於低菌濃度時，流道內即時檢測之方式則會因菌體易吸附在管壁上而效果降低。

Halobacterium salinarum contains purple membranes (PM) constituted of a light-sensitive protein, bacteriorhodopsin, which generates a proton gradient across the membrane upon illumination. Therefore, PM can be used to produce photocurrents. Based on the light scattering ability of bacteria and the linear correlation between PM photocurrents and illumination intensities, this study demonstrated a sensitive PM-based biochip to quantitatively detect Streptococcus mutans. Two kinds of biochips were developed, each fabricated with different recognition molecules on PM-coated chips, covalently conjugated S. mutans-specific aptamers and anti-S. mutans antibodies orientationally immobilized through covalently linked protein A. Both microfluidic real-time and static detections of S. mutans using either chip were demonstrated and the detections were evidenced by both fluorescence microscopy and photocurrent reduction analyses of the cell-bound chips. For the static detection, the antibody-PM composite chip exhibited the sensitivity of 10 CFU/mL, while the aptamer-PM one had the sensitivity further down to 1 CFU/mL. Both chips could detect 106 CFU/mL cultures in a single step without prior dilution. S. mutans cultures at high concentrations were real-time detected in the microfluidic device as accurately and effectively as in a cuvette, while the detection of very low-populated cultures by microfluidics was poor due to cell adsorption on microfluidic device and tubing surfaces.

吳欣穎, 細菌視紫質泛用型免疫光電晶片製備與原子力顯微鏡分析, 國立台灣科技大學化學工程研究所碩士論文 2015

陳冠辰, 適體-細菌視紫質生物光電感測晶片之探討, 國立台灣科技大學化學工程研究所碩士論文 2015

廖信詮, 微流體於細菌視紫質光電晶片製備之應用, 國立台灣科技大學化學工程研究所碩士論文 2015

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