(3.238.36.32) 您好!臺灣時間:2021/02/27 08:51
字體大小: 字級放大   字級縮小   預設字形  
回查詢結果

詳目顯示:::

我願授權國圖
: 
twitterline
研究生:許偉恩
研究生(外文):Wei-En Hsu
論文名稱:修飾低吸附雙極性多胜肽金表面於胎盤生長因子檢測應用
論文名稱(外文):Surface Functionalization of Gold Surfaces with Polypeptide: A Low-Fouling Zwitterionic Surface for Detect Placenta Growth Factor
指導教授:林啟萬林啟萬引用關係
口試委員:施文彬彭盛裕魏淑鉁
口試日期:2017-06-09
學位類別:碩士
校院名稱:國立臺灣大學
系所名稱:醫學工程學研究所
學門:工程學門
學類:綜合工程學類
論文種類:學術論文
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:表面電漿子共振抗吸附兩性離子大腸直腸癌胎盤生長因子
外文關鍵詞:Surface Plasmon ResonanceAnti-foulingZwitterionColorectal CancerPlacental Growth Factor
相關次數:
  • 被引用被引用:0
  • 點閱點閱:90
  • 評分評分:系統版面圖檔系統版面圖檔系統版面圖檔系統版面圖檔系統版面圖檔
  • 下載下載:0
  • 收藏至我的研究室書目清單書目收藏:0
在台灣大腸直腸癌的發生已經率逐年提高,目前大腸直腸癌是全台灣所有癌症中發生率第一名的癌症,已經七度蟬聯癌症發生人數的第一名,且大腸直腸癌在死亡率上也高居第三,僅次於肝癌和肺癌的死亡率。根據過去的許多癌症研究中發現,目前對於大腸直腸癌的初期診斷,胎盤生長因子(PlGF)是一個重要的指標,可以在患病初期及早發現並治療,並且胎盤生長因子也可以當成預後復發的危險因子指標之一。
本研究為了能夠檢測胎盤生長因子的存在,以表面電漿子共振技術來進行量測,該技術具有高靈敏度、即時量測且不須螢光標記等優點,並且使用抗體為感測元件,能夠對於標的物有良好的結合能力。但由於胎盤生長因子是存在於血清中,血清檢體包含了大量的蛋白質,在接觸量測表面時會受到極大的非特異性吸附影響,導致錯誤的訊號產生。為了解決這個現象,本研究使用混合的胺基酸序列來進行蛋白質的抗吸附處理與抗體的固定化,以簡單的方式達到保護量測表面的同時,也可以用於檢測胎盤生長因子的存在。在溶液中,本研究所提出的量測方法已經可以有效的檢測出PlGF的存在,且檢測極限可以到達2pg/ml。在實際臨床樣本的量測中,由於非特異性吸附的影響,目前與現有ELISA技術相比,同樣可以擁有六成的精確度。
In Taiwan, the incidence of colorectal cancer has been increasing year by year. Colorectal cancer is the first occurrence of all cancer. The incidence rate of colorectal cancer has become first for seven consecutive years and the mortality rate also ranked third, only behind liver cancer and lung cancer. According to many cancer studies in the past, placental growth factor(PlGF) is the important marker that can help us to know the potential patient and give them some treatment. In addition to this, PlGF can also be used as one of the risk factors for prognosis recurrence.
In this study, I want to know if PlGF exists in patient''s serum. I used surface plasmon resonance(SPR) technology for detection of PlGF. The reasons being that SPR has many advantages, like high sensitivity, real-time detection, non-Fluorescent label and so on. Moreover, I used the antibody as my sensing element. It has a good ability to combine with the target.
However, PlGF exists in the serum contains a large amount of protein and will be affected by non-specific adsorption, therefore resulted in the error signal. In order to solve this problem, I used the mixed amino acid sequence for protein anti-fouling and antibody modification in this study. This simple method can achieve protection of surface, and also can be used to detect if PlGF exists at the same time. In the solution, the measurement method proposed in this study has been able to effectively detect the presence of PlGF, and the limit of detection(LOD) is 2pg/ml. Due to the impact of non-specific adsorption, if compare the actual clinical sample with the existing ELISA technology, this study also have sixty precent accuracy.
口試委員會審定書 I
致謝 II
中文摘要 III
ABSTRACT IV
目錄 V
圖目錄 VII
表目錄 IX
第 1 章 緒論 1
1.1 前言 1
1.2 研究動機 3
1.3 論文架構 4
第 2 章 文獻回顧 5
2.1 生物感測器在癌症上的發展 5
2.2 癌症的偵測與診斷 5
2.2.1 生物標記物 5
2.2.2 大腸直腸癌預後診斷 6
2.2.3 大腸直腸癌腫瘤標記物 8
2.3 表面電漿子共振(SPR) 10
2.3.1 表面電漿子共振原理 10
2.3.2 表面電漿子激發方式 12
2.3.3 表面電漿子共振量測方式 15
2.4 抵抗蛋白質的非特異性吸附 17
2.4.1 抵抗蛋白質非特異性吸附之材料 17
2.4.2 以氨基酸組成之兩性離子材料 19
2.5 抗體於生物感測器表面的修飾 21
2.5.1 抗體與感測元件表面 21
2.5.2 修飾抗體之方向性 23
2.5.3 多胜肽材料應用於抗體的方向性修飾 25
第 3 章 研究材料與方法 26
3.1 表面電漿子共振量測系統架構 26
3.2 軟體量測介面 30
3.3 晶片製程 32
3.4 抗蛋白質吸附材料與抗體的修飾方法 35
3.5 量測訊號處理 38
第 4 章 研究結果與討論 39
4.1 抗蛋白質吸附層 39
4.1.1 不同抗吸附材料對於非特異性吸附之效果 39
4.1.2 抗吸附層修飾之最佳濃度 42
4.2 抗蛋白質吸附材料與大腸直腸癌抗體修飾 43
4.2.1 不同比例之抗吸附層與抗體修飾測試 43
4.3 大腸直腸癌臨床測試結果 45
4.3.1 以胎盤生長因子標準品驗證抗體鍵結效果 45
4.3.2 以表面電漿子共振檢測血清中的胎盤生長因子 46
第 5 章 結論與未來展望 52
第 6 章 參考文獻 54
第 7 章 附錄 59
7.1 抗吸附材料測試之SRP感測圖 59
7.2 臨床血清樣本測試之SPR感測圖 61
7.2.1 大腸癌病人血清樣本 61
7.2.2 對照組血清樣本 62
[1] Cancer Council NSW, “Understanding Cancer – What is cancer?” 民國106年4月21號,取自: http://languages.cancercouncil.com.au/zh-hant/
[2] 林鵬展醫師、陳立宗醫師,“大腸直腸癌(Colorectal Cancer)”,財團法人台灣癌症基金會,民國106年5月1,取自: http://www.canceraway.org.tw/
[3] Hanahan, Douglas, and Robert A. Weinberg. "The hallmarks of cancer." cell 100.1 (2000): 57-70.
[4] Dewerchin, Mieke, and Peter Carmeliet. "Placental growth factor in cancer." Expert opinion on therapeutic targets 18.11 (2014): 1339-1354.
[5] Rasooly, Avraham, and James Jacobson. "Development of biosensors for cancer clinical testing." Biosensors and Bioelectronics 21.10 (2006): 1851-1858.
[6] Burt, Randall W., et al. "Colorectal cancer screening." Journal of the National Comprehensive Cancer Network 8.1 (2010): 8-61.
[7] Bethesda, M., SEER Cancer Statistics Factsheets: Colon and Rectum Cancer. National Cancer Institute. http://seer.cancer.gov/statfacts/html/colorect.html.
[8] Siegel, Rebecca L., Kimberly D. Miller, and Ahmedin Jemal. "Cancer statistics, 2016." CA: a cancer journal for clinicians 66.1 (2016): 7-30.
[9] Ludwig, Joseph A., and John N. Weinstein. "Biomarkers in cancer staging, prognosis and treatment selection." Nature Reviews Cancer 5.11 (2005): 845-856.
[10] Wei, Shu-Chen, et al. "Preoperative serum placenta growth factor level is a prognostic biomarker in colorectal cancer." Diseases of the Colon & Rectum 52.9 (2009): 1630-1636.
[11] Homola, Jiří. "Electromagnetic theory of surface plasmons." Surface plasmon resonance based sensors. Springer Berlin Heidelberg, 2006. 3-44.
[12] 邱國斌, and 蔡定平. "金屬表面電漿簡介." 物理雙月刊 28.2 (2006): 472-485.
[13] Nelson, S. G., Kyle S. Johnston, and Sinclair S. Yee. "High sensitivity surface plasmon resonace sensor based on phase detection." Sensors and actuators B: Chemical 35.1-3 (1996): 187-191.
[14] Kabashin, A. V., and P. I. Nikitin. "Surface plasmon resonance interferometer for bio-and chemical-sensors." Optics communications 150.1 (1998): 5-8.
[15] Micallef, C., P. Cuschieri, and M. R. Bonnici. "Contamination of contact-lens-related sources with Pseudomonas aeruginosa." Ophthalmologica 214.5 (2000): 324-331.
[16] Brash, John L. "Exploiting the current paradigm of blood–material interactions for the rational design of blood-compatible materials." Journal of Biomaterials Science, Polymer Edition 11.11 (2000): 1135-1146.
[17] Hatakeyama, Evan S., et al. "New protein-resistant coatings for water filtration membranes based on quaternary ammonium and phosphonium polymers." Journal of membrane science 330.1 (2009): 104-116.
[18] Briand, Jean-Francois. "Marine antifouling laboratory bioassays: an overview of their diversity." Biofouling 25.4 (2009): 297-311.
[19] 馬春風. "抗蛋白吸附聚合物的合成與性質. " 中國科學技術大學, 2011.
[20] Amanda, Aurelia, and Surya K. Mallapragada. "Comparison of Protein Fouling on Heat‐Treated Poly (vinyl alcohol), Poly (ether sulfone) and Regenerated Cellulose Membranes Using Diffuse Reflectance Infrared Fourier Transform Spectroscopy." Biotechnology progress 17.5 (2001): 917-923.
[21] Johnell, Matilda, Rolf Larsson, and Agneta Siegbahn. "The influence of different heparin surface concentrations and antithrombin-binding capacity on inflammation and coagulation." Biomaterials 26.14 (2005): 1731-1739.
[22] McArthur, Sally L., et al. "Effect of polysaccharide structure on protein adsorption." Colloids and Surfaces B: Biointerfaces 17.1 (2000): 37-48.
[23] Chen, Shengfu, et al. "Strong resistance of phosphorylcholine self-assembled monolayers to protein adsorption: insights into nonfouling properties of zwitterionic materials." Journal of the American Chemical Society 127.41 (2005): 14473-14478.
[24] Holmlin, R. Erik, et al. "Zwitterionic SAMs that resist nonspecific adsorption of protein from aqueous buffer." Langmuir 17.9 (2001): 2841-2850.
[25] Prime, Kevin L., and George M. Whitesides. "Adsorption of proteins onto surfaces containing end-attached oligo (ethylene oxide): a model system using self-assembled monolayers." Journal of the American Chemical Society 115.23 (1993): 10714-10721.
[26] Roberts, Carmichael, et al. "Using mixed self-assembled monolayers presenting RGD and (EG) 3OH groups to characterize long-term attachment of bovine capillary endothelial cells to surfaces." Journal of the American Chemical Society 120.26 (1998): 6548-6555.
[27] Golander, C. G., et al. "Poly (ethylene glycol) chemistry: biotechnical and biomedical applications." (1992).
[28] Rabinow, B. E., et al. "Biomaterials with permanent hydrophilic surfaces and low protein adsorption properties." Journal of Biomaterials Science, Polymer Edition 6.1 (1995): 91-109.
[29] He, Yi, et al. "Molecular simulation studies of protein interactions with zwitterionic phosphorylcholine self-assembled monolayers in the presence of water." Langmuir 24.18 (2008): 10358-10364.
[30] Ishihara, Kazuhiko, et al. "Why do phospholipid polymers reduce protein adsorption?." Journal of biomedical materials research 39.2 (1998): 323-330.
[31] Chen, Shengfu, Lingyun Liu, and Shaoyi Jiang. "Strong resistance of oligo (phosphorylcholine) self-assembled monolayers to protein adsorption." Langmuir 22.6 (2006): 2418-2421.
[32] Yaseen, M., et al. "The structure of zwitterionic phosphocholine surfactant monolayers." Langmuir 22.13 (2006): 5825-5832.
[33] Zheng, Jie, et al. "Strong repulsive forces between protein and oligo (ethylene glycol) self-assembled monolayers: A molecular simulation study." Biophysical journal 89.1 (2005): 158-166.
[34] Bolduc, Olivier R., Joelle N. Pelletier, and Jean-François Masson. "SPR biosensing in crude serum using ultralow fouling binary patterned peptide SAM." Analytical chemistry 82.9 (2010): 3699-3706.
[35] Lin, Peter, et al. "Nonfouling property of zwitterionic cysteine surface." Langmuir 30.22 (2014): 6497-6507.
[36] Nowinski, Ann K., et al. "Sequence, structure, and function of peptide self-assembled monolayers." Journal of the American Chemical Society 134.13 (2012): 6000-6005.
[37] Engvall, Eva, and Peter Perlmann. "Enzyme-linked immunosorbent assay (ELISA) quantitative assay of immunoglobulin G." Immunochemistry 8.9 (1971): 871-874.
[38] Le Brun, Anton P., et al. "The structural orientation of antibody layers bound to engineered biosensor surfaces." Biomaterials 32.12 (2011): 3303-3311.
[39] Trilling, Anke K., Jules Beekwilder, and Han Zuilhof. "Antibody orientation on biosensor surfaces: a minireview." Analyst 138.6 (2013): 1619-1627.
[40] Kriech, Douglas Michael. Single molecule fluorescence imaging of protein-surface and protein-protein interactions at a glass-water interface. The University of Utah, 2015. [41] Tajima, Nobuyuki, Madoka Takai, and Kazuhiko Ishihara. "Significance of antibody orientation unraveled: well-oriented antibodies recorded high binding affinity." Analytical chemistry 83.6 (2011): 1969-1976.
[42] Liu, Fang, et al. "Immobilized antibody orientation analysis using secondary ion mass spectrometry and fluorescence imaging of affinity-generated patterns." Analytical chemistry 82.7 (2010): 2947-2958.
[43] Karyakin, Arkady A., et al. "Oriented immobilization of antibodies onto the gold surfaces via their native thiol groups." Analytical chemistry 72.16 (2000): 3805-3811.
[44] Kausaite-Minkstimiene, A., et al. "Comparative study of random and oriented antibody immobilization techniques on the binding capacity of immunosensor." Analytical chemistry 82.15 (2010): 6401-6408.
[45] Boozer, Christina, et al. "DNA directed protein immobilization on mixed ssDNA/oligo (ethylene glycol) self-assembled monolayers for sensitive biosensors." Analytical chemistry 76.23 (2004): 6967-6972.
[46] Vijayendran, Ravi A., and Deborah E. Leckband. "A quantitative assessment of heterogeneity for surface-immobilized proteins." Analytical Chemistry 73.3 (2001): 471-480.
[47] Song, Hong Yan, et al. "Comparative study of random and oriented antibody immobilization as measured by dual polarization interferometry and surface plasmon resonance spectroscopy." Langmuir 28.1 (2011): 997-1004.
[48] Iijima, Masumi, et al. "Nano-visualization of oriented-immobilized IgGs on immunosensors by high-speed atomic force microscopy." Scientific reports 2 (2012).
[49] Jung, Yongwon, et al. "Controlled antibody immobilization onto immunoanalytical platforms by synthetic peptide." Analytical biochemistry 374.1 (2008): 99-105.
[50] Tsai, Ching-Wei, et al. "Strategy of Fc-recognizable peptide ligand design for oriented immobilization of antibody." Analytical chemistry 86.6 (2014): 2931-2938.
QRCODE
 
 
 
 
 
                                                                                                                                                                                                                                                                                                                                                                                                               
第一頁 上一頁 下一頁 最後一頁 top
系統版面圖檔 系統版面圖檔