研究生: |
林聖為 Sheng-wei Lin |
---|---|
論文名稱: |
利用自排性單分子膜與生物親和吸附增進PM晶片的製程 Coupling of self-assembled monolayer and bioaffinity adsorption to improve purple-membrane chip fabrication |
指導教授: |
陳秀美
Hsiu-mei Chen |
口試委員: |
戴 龑
Yian Tai 陳良益 Liang-yih Chen |
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: | 2012 |
畢業學年度: | 100 |
語文別: | 中文 |
論文頁數: | 130 |
中文關鍵詞: | 自排性單分子膜 、細菌視紫質 、紫膜 、木瓜蛋白酶 |
外文關鍵詞: | SAM, bacteriorhodopsin, purple membrance, papain |
相關次數: | 點閱:187 下載:0 |
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存在於Halobacterium salinarium紫膜 ( Purple membrance,PM ) 內的bacteriorhodopsin 蛋白質受光後可產生光電響應。本研究嘗試使用不同自排性單分子膜( self-assembled monolayer,SAM )分子以及不同PM固定法,試圖得到最佳的PM晶片製程以及控制PM的塗佈方向。所研究不同碳鏈長度的羧亞磷酸SAM分子分別為16PHDA、11PUA、6PHA以及3PPA,其中以16PHDA修飾ITO的表面最為疏水,接觸角約為90度上下,11PHA為60-70度,6PHA為20-30度,3PPA則為10-20度。在CV分析中,16PHDA氧化峰電流值為-37μA,還原峰電流值為34μA,這兩個數值大小會隨著所用SAM碳鏈的減少而上升,到3PPA時氧化峰電流值為-43μA,還原峰電流值為43μA。此外,生物親和吸附固定化的製程,以AFM分析發現PM塗佈於以長鏈SAM修飾的ITO表面時會彎曲拱起,使晶片擁有較高的微分光電流值以及較大的瞬態脈衝光電流訊號B2 的衰減時間。極性高的溶劑有利於6PHA以亞磷酸端貼附於ITO上,PM在胺磷酸SAM APPA上的塗佈量優於3PPA。此外也發現ITO之表面與粗糙度會影響PM的貼覆以及微分電流響應速度。再者,另以單純的共價鍵結進行PM固定化,發現不論使用懸浮在pH 4.5、5.2或者7的PM或水解PM,兩者都是以胞內側定向於被16PHDA修飾過之ITO上,在4.5與7這兩個pH值下,被水解的PM其光電流會比未水解的PM來的高;而在pH 5.2時則得到相反的結果。最後,比較生物親和吸附與共價鍵結固定化之製程,可發現前者可得到比後者更高的PM塗佈量,但兩者PM皆是以胞內側定向於被SAM修飾之ITO。另一方面,以AFM分析可發現,被共價鍵結固定化的PM較為平整,而以生物親和吸附法固定化的PM其形態則較為彎曲拱起。
Bacteriorhodopsin residing in the purple membrane (PM) of Halobacterium salinarum can generate photoelectric responses after being illuminated. In this study, different self-assembled monolayer (SAM) molecules as well as immobilization methods were employed to optimize the fabrication process of PM chips and to control the orientation of immobilized PMs. The contact angles of ITO varied with the lengths of coating carboxyalkylphosphonic SAMs, with 16PHDA yielding the most hydrophobic surface (∼90°) and 3PPA the most hydrophilic one (10-20°). In CV analyses, the oxidation and reduction peak currents of 16PHDA-coated ITO were -37 and 34 μA, respectively, with both values increasing with the SAM length and reaching to -43 and 43μA, respectively, for 3PPA. In addition, PM bioaffinity-immobilized on longer carboxyalkylphosphonic SAM-coated ITO was wedge-shaped in appearance as revealed by AFM analyses, yielding higher differential photocurrents and longer decay time of the B2 pulse photocurrents than the PM coated on shorter SAMs. Polar solvents favored the adsorption of 6PHA on ITO through its phosphonic end, and the coating amount of PM on an aminophosphonic SAM, APPA, was higher than that on its length-equivalent carboxyalkylphosphonic SAM, 3PPA. Furthermore, the surface morphology and roughness of ITO had effects on the coating and differential photocurrent response rate of coated PM. Moreover, PM covalently immobilized on 16PHDA-coated ITO was orientated with its cytoplasmic side facing ITO no matter the coating conditions. The papain-digested PM generated higher photocurrents than the undigested one when coated with pH 4.5 and 7 suspension buffers, while the results were vice versa when a pH 5.2 buffer was used. Compared to affinity immobilization, the covalent method led to a much less coating amount and more stretching of PM; however, both methods yielded the same PM orientation.
陳逸航,"定向性細菌視紫質晶片之光電與二倍頻響應探討",國立台灣科技大學化學工程研究所碩士論文 (2010)
蔡孟訓,"自排性單層分子膜於Bacteriorhodopsin光電晶片製備之探討",國立台灣科技大學化學工程研究所碩士論文 (2011)
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