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Please use this identifier to cite or link to this item: http://ntour.ntou.edu.tw:8080/ir/handle/987654321/34654

Title: 利用生物功能性金奈米材料於細菌檢測
Detection of Bacteria by Using Biofunctional Gold Nanomaterials
Authors: Yu-Ting Tseng
Contributors: NTOU:Institute of Bioscience and Biotechnology
Keywords: 金奈米粒子;金奈米量子點;螢光;細菌;雷射游離脫附質譜儀;硝化纖維膜
Gold Nanoparticle;Gold nanodots;Luminescent;Bacteria;Laser desorption/ionization mass spectrometer;Nitrocellulose membrane
Date: 2013
Issue Date: 2013-10-07T02:44:52Z
Abstract: 本研究主要開發生物功能性金奈米材料(gold nanomaterials),應用於偵測並抑制細菌生長。(1)我們利用修飾甘露糖(mannose)之金奈米量子點(Man–Au NDs)之螢光,開發出一套簡單且方便的檢測系統。利用Man–Au NDs與大腸桿菌(Escherichia coli;E. coli)表面線毛上FimH蛋白之甘露糖受體,具有專一性鍵結,可以使Man–Au NDs與E. coli結合,藉此能夠定量E. coli。利用偵測結合上E. coli之Man–Au NDs螢光,可檢測E. coli最低偵測極限為150 CFU/mL,另外Man–Au NDs也可以被視為抗菌劑,因Man–Au NDs能夠選擇性使E. coli團聚,進而造成抑制細菌生長的效果,同時由於Man–Au NDs表面修飾官能基十分密集,故可以應用於真實樣品中,而不會受到樣品中基質的干擾。(2) 我們開發出利用基質輔助雷射游離脫附質譜儀(matrix-assisted laser desorption/ionization mass spectrometer;MALDI-MS)搭配上表面修飾抗體(antibody)金奈米粒子(AbAu NPs)之感測器,AbAu NPs是由5.5-nm金奈米粒子和能與細菌專一性結合之抗體所組成,將AbAu NPs與樣品中細菌結合後,以離心純化AbAu NPs/細菌之複合物,並將其點樣在硝化纖維膜(nitrocellulose membrane;NCM;0.3 cm×0.3 cm;孔徑: 0.45 μm)上進MALDI-MS做偵測,我們可以利用金離子團簇(Au clusters)的訊號,進而推測目標細菌濃度。此AbAu NPs/NCM探針,具有高靈敏度,其最低偵測極限為500 CFU/mL,且改變修飾於金奈米粒子之抗體,即可以選擇性偵測E. coli、金黃色葡萄球菌(Staphylococcus aureus; S. aureus)或沙門氏菌(Salmonella enterica;S. enterica)等不同細菌,並能於自來水、果汁、牛奶和尿液等真實樣品中做偵測。
In our article, we demonstrated biofunctional gold nanomaterials could be developed for detection and inhibition of the growth of bacteria, respectively. (1)We developed a simple and convenient method for synthesis of luminescent mannosed-capped gold nanodots (Man–Au NDs). Multivalent interactions between the Man–Au NDs and FimH proteins located on the bacterial pili of Escherichia coli (E. coli) resulted in the formation of aggregated bacterial clusters. Monitoring the luminescence of the Man–Au NDs allowed the detection of E. coli at levels as low as 150 CFU/mL. The Man–Au NDs were also an efficient antibacterial agent, selectively inhibiting the growth of E. coli through Man–Au ND–induced agglutination. Our small-diameter Man–Au NDs, which provided an ultrahigh ligand density (local concentration) of mannose units for antibacterial agent in real samples. (2)We described a novel method for bacterial detection using mass spectrometry and antibody-conjugated gold nanoparticles (AbAu NPs). The AbAu NPs were obtained from as-prepared 5.5 nm Au NPs and bacteria-specific antibodies. After binding of the AbAu NPs to the bacteria, the purified AbAu NPs/bacteria conjugates were deposited on a nitrocellulose membrane (NCM; 0.3 cm×0.3 cm; pore size: 0.45 μm) prior to LDI-MS analysis. The Au ion signals from the Au NPs using LDI were employed to analyze bacteria. Under optimal conditions, this AbAu NPs/NCM probe is highly sensitive [limit of detection ~500 colony forming units (CFU)/mL] and selective toward bacteria E. coli, Staphylococcus aureus (S. aureus), or Salmonella enterica (S. enterica) by using the corresponding Ab–Au NPs. Therefore, this LDI-MS approach provides a simple and cost-effective route for the high-speed analysis of low concentrations of bacteria with ultrahigh sensitivity and selectivity.
URI: http://ethesys.lib.ntou.edu.tw/cdrfb3/record/#G0010036030
Appears in Collections:[生命科學暨生物科技學系] 博碩士論文

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