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Enhancing sensitivities of immunoassays by nanoenvironment
|關鍵字:||3D gold nano particle seneor;3D奈米金顆粒感測器;serpentine microfludic channel;Der p2;S型微流裝置;歐洲式塵螨||出版社:||機械工程學系所||引用:|| Thomas WR, Smith WA, Hales BJ, Mills KL, O''Brien RM: Characterizationand immunobiology of house dust mite allergens. Int Arch Allergy Immunol 2002, 129(1):1-18.  J. J. Tsai, H. D. Shen, K. Y. Chua, Purification of group 2 Dermatophagoides pteronyssinus allergen and prevalence of its specific IgE in asthmatics, Int Arch Allergy Immunol, 121(3), 205-210, 2000.  J. J. Tsai, J. Y. Yen, Y. H. Yang, The Prevalence of Der p 2 Allergy in Asthmatic Patients in Taiwan. 2nd Congress of the Federation of Immunological Societies of Asia-Oceania, 129-35, 2000.  E. C. Liao, E. L. Hsu, J. J. Tsai, C. M. Ho, Immunologic Characterization and Allergenicity of Recombinant Tyr p 3 Allergen from the Storage Mite Tyrophagus putrescentiae, Int Arch Allergy Asthma Innumol. (IN press)  E. C. Liao, C. M. Ho, J. J. 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A 3D gold nano particle (NP) seneor in serpentine microfludic channel (S channel) was developed and has the advantage of label-free detection, high sensitivity and low cost. In the fabrication of 3D gold NP sensor, we demonstrated a replica mold fabrication method for nano hemisphere arrays by nanomolding nickel electroforming the barnier-layer surface of an anodic aluminum oxide (AAO) as the master mold. The fabricated 3D nickel mold is further used for replica molding of a nano-structure polycarbonate (PC) substrate by hot embossing. Afterwards, a layer of gold film and 3D NPs were deposited on the PC substrate by sputtering and electrochemical deposition, respectively. Compared to a flat gold film, 3D gold NPs have 3 times surface area which can highly increase the sensitivity of sensor. Furthermore, a serpentine microfludic channel was fabricated by soft lithography and polydimethylsiloxane (PDMS). Then, 3D gold NPs sensor were bonded on the S channel by O2 plasma treatment. D. pteronyssinus (Der p2) and specific antibody (C1) were used to present the performance of sensor. Electrochemical impedance spectroscopy (EIS) and fluorescence detection by confocal microscope were used to detect the concentration of Der p2.
In the EIS analysis, the minimum detectable concentrations of Der p2 were down to 0.1pg/ml and 10pg/ml when the 3D gold NPs sensor in S channel and in immerse deposition, respectively. In the fluorescence detection, the average coverages were 35.59% and 11.44% at the concentration of Der p2 10pg/ml, when sensor in S channel and in immerse deposition. Experiment at results showed that 3D gold NP sensor in S channel have 100 times sensitivity than sensor than sensor in immerse deposition. In the other hand, EIS analysis showed larger sensing linearity than fluorescence detection because fluorescence was saturated when the concentration of Der p2 was larger than 1ng/ml. Our results showed that 3D gold NPs sensor in S channel have great sensitivity (0.1pg/ml) compared to ELISA (1ng/ml).
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