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研究生: 林宸宇
Lin, Chen-Yu
論文名稱: 水溶性碳量子點的合成及其應用
The characterizations and applications of water-soluble carbon quantum dots
指導教授: 葉怡均
Yeh, Yi-Chun
學位類別: 碩士
Master
系所名稱: 化學系
Department of Chemistry
論文出版年: 2017
畢業學年度: 105
語文別: 中文
論文頁數: 63
中文關鍵詞: 碳量子點生物相容性細胞標記Pb5FhuA細胞裂解
英文關鍵詞: carbon quantum dots, biocompatibility, bioimaging, Pb5, FhuA, cell lysis
DOI URL: https://doi.org/10.6345/NTNU202202358
論文種類: 學術論文
相關次數: 點閱:111下載:11
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  • 碳量子點是一種新型的零維奈米材料,由於普遍具有良好的螢光特性和生物相容性,在生物顯像的領域受到廣泛的應用。本研究我們以檸檬酸和三乙烯四胺為前驅物,並利用微波法合成氮摻雜碳量子點。在應用方面,第一,不同的酸鹼環境影響下,其螢光強度隨著pH值有良好的線性關係;第二,針對不同的金屬離子溶液,發現對六價鉻離子有明顯的專一性螢光消逝;第三,我們也透過碳量子點做大腸桿菌的細胞標記,發現有良好的螢光強度和生物相容性。我們利用大腸桿菌BL21將Pb5蛋白大量表現純化後以螢光基團修飾,藉由噬菌體Pb5蛋白和大腸桿菌外膜蛋白的結合特性,作為標記大腸桿菌的方式,也發現對於大量表現FhuA外膜蛋白的大腸桿菌可作為良好的專一性螢光探針。再者,我們能以螢光訊號的強度來呈現不同細菌量對於修飾過的Pb5蛋白結合情況,發現有良好的線性關係。最後,我們利用偵測在細胞裂解產物的上清液螢光訊號,定量細胞裂解的效率。

    Carbon quantum dot is a type of zero-dimensional nanomaterials. These materials have been widely applied to bioimaging and sensor to their outstanding properties such as fluorescence and biocompatibility. In this study, water-soluble N-doped carbon quantum dots (CQDs) were synthesized by microwave-induced decomposition using citric acid and triethylenetetramine (TETA) as precursors. For the applications, firstly, the intensity of fluorescence showed linear relationship with pH values. Secondly, these CQDs were sensitive fluorescent probes for the detection of Cr6+ with significant fluorescence quenching. Thirdly, we found the excellent properties of cell imaging and biocompatibility on labeling E. coli with CQDs. We labeled the E. coli outer membrane with the strong interactions between the bacteriophage tail protein Pb5 which was modified with rhodamine and E. coli outer membrane protein ferric hydroxamate uptake A (FhuA). Besides, we could estimate the number of the cells based on the linear relationship between the fluorescence signals of modified Pb5 and the cells. Finally, the protein complexes, FhuA-Pb5-rhodamine, was used to estimate the efficiency of cell lysis.

    Chapter I 緒論 1 1.1 奈米材料的特性 1 1.1.1 奈米材料的維度 1 1.1.2 表面積效應 2 1.1.3 量子侷限效應 2 1.2 碳量子點 3 1.2.1 簡介 3 1.2.2 碳量子點的製備方法 4 1.2.3 碳量子點的應用與發展 7 1.3 噬菌體 9 1.3.1 簡介 9 1.3.2 分類 9 1.3.3 T-type 噬菌體 10 1.3.4 感染與生長 10 1.3.5 烈性噬菌體與溫和性噬菌體 12 1.4 大腸桿菌鐵離子通道蛋白 13 1.4.1 鐵載體 13 1.4.2 鐵離子通道蛋白FhuA 15 1.5 噬菌體Pb5蛋白和E. coli FhuA蛋白的結合 17 1.6 細胞顯像標記 18 1.7 細胞裂解 19 1.8 研究動機與目標 20 Chapter II 實驗 21 2.1 實驗藥品 21 2.2 實驗儀器 23 2.3 實驗設計 23 2.4 實驗方法 24 2.4.1 實驗器材的處理 24 2.4.2 碳量子點的製備 24 2.4.3 合成條件的最佳化 24 2.4.4 穩定性 25 2.4.5 酸鹼環境的影響 25 2.4.6 金屬離子偵測的專一性 26 2.4.7 碳量子點對Cr6+的專一性 26 2.4.8 金屬離子偵測的干擾性 26 2.4.9 噬菌體Pb5蛋白的純化 26 2.4.10 蛋白質電泳 29 2.4.11 Pb5的修飾 32 2.4.12 Pb5-rhodamine和其他菌種的結合性測試 32 2.4.13 以螢光顯微鏡觀察Pb5-rhodamine和其他菌種的結合 33 2.4.14 Pb5-rhodamine和單位菌量的關係 33 2.4.15 以FhuA-Pb5-rhodamine來呈現細胞裂解程度 35 Chapter III 實驗結果與討論 36 3.1 碳量子點的光學特性與合成條件的最佳化 36 3.1.1 光學特性 36 3.1.2 前驅物濃度對螢光的影響 37 3.1.3 加熱時間對螢光的影響 38 3.2 碳量子點的穩定性 39 3.3 碳量子點的性質 40 3.3.1 X射線繞射圖 40 3.3.2 X射線電子能譜圖 41 3.3.3 穿隧式電子顯微鏡影像 42 3.4 碳量子點對酸鹼值的螢光變化 43 3.4.1 不同酸鹼環境 43 3.4.2 可逆性 45 3.5 碳量子點對金屬離子的專一性和干擾性測試 46 3.5.1 專一性 46 3.5.2 六價鉻離子濃度與螢光的關係 47 3.5.3 干擾性 48 3.6 碳量子點對大腸桿菌的標記 49 3.7 噬菌體Pb5蛋白的純化 50 3.8 蛋白質複體Pb5-rhodamine的應用 51 3.8.1 不同菌種的標記 51 3.8.2 不同菌種的標記-螢光顯微鏡 53 3.8.3 與細菌數量的關係 54 3.8.4 判斷細胞裂解效果 55 Chapter IV 實驗結果討論與結論 57 4.1 探討碳量子點在不同酸鹼環境的螢光變化 57 4.2 探討碳量子點於金屬離子存在下的螢光變化 59 4.3 結論 60 參考文獻 61

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