A surface plasmon resonance immunosensor for detecting a dioxin precursor using a gold binding polypeptide |
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Authors: | Soh Nobuaki Tokuda Tomoyuki Watanabe Tomomi Mishima Keiko Imato Toshihiko Masadome Takashi Asano Yasukazu Okutani Saeko Niwa Osamu Brown Stanley |
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Institution: | a Department of Chemical Systems and Engineering, Graduate School of Engineering, Kyushu University, Hakozaki, Higashi-ku, Fukuoka 812-8581, Japan b Department of Chemical Science and Engineering, Ariake National College of Technology, Omuta, Fukuoka 836-8585, Japan c Department of Chemical and Biological Engineering, Hachinohe National College of Technology, Hachinohe, Aomori 039-1192, Japan d NTT Microsystem Integration Laboratories, Morinosato, Wakamiya, Atsugi-shi, Kanagawa 243-0198, Japan e Department of Molecular Cell Biology, University of Copenhagen, Øster Farimagsgade 2A, DK-1353 Copenhagen K, Denmark |
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Abstract: | A surface plasmon resonance (SPR) based biosensor was developed for monitoring 2,4-dichlorophenol, a known dioxin precursor, using an indirect competitive immunoassay. The SPR sensor was fabricated by immobilizing a gold-thin layer on the surface of an SPR sensor chip with an anti-(2,4-dichlorophenol) antibody using a gold binding polypeptide (GBP) and protein G. The SPR response based on the antigen-antibody reaction in a flow system was measured by injecting a 2,4-dichlorophenol sample solution into the flow system in which the SPR sensor was located. In a direct immunoassay system using the modified sensor chip, no significant SPR angle shift less than 0.001° was observed when a 25 ppm of 2,4-dichlorophenol solution was injected. In order to improve the sensitivity of the SPR sensor, an indirect competitive immunoassay method was used in conjunction with the SPR sensor system using 2,4-dichlorophenol conjugated with bovine serum albumin (BSA). In the competitive assay, a 350 ppm 2,4-dichlorophenol-BSA conjugate solution containing 2,4-dichlorophenol at various concentrations (10-250 ppb) were injected into the SPR sensor system. The sensitivity of this indirect immunoassay was found to be extremely sensitive, compared to the direct one, and a detection limit of 20 ppb was estimated. Verification that the use of GBP for immobilizing the antibody on the sensor chip enhanced the sensitivity to 2,4-dichlorophenol was obtained by comparing the procedure with another modification, in which BSA was used instead of GBP for immobilizing the antibody on the sensor chip. The affinity constant of 2,4-dichlorophenol and its conjugate to the antibody were estimated form the SPR response. |
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Keywords: | SPR sensor 2 4-Dichlorophenol Dioxins Endocrine disrupting chemicals Gold binding polypeptide Protein G |
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