Analytical Sciences

Flow Immunoassay for Nonioinic Surfactants Based on Surface Plasmon Resonance Sensors

Mayumi TANAKA,*,** Kazuhira SAKAMOTO,** Hizuru NAKAJIMA,* Nobuaki SOH,* Koji NAKANO,* Takashi MASADOME,*** and Toshihiko IMATO*

*Department of Applied Chemistry, Graduate School of Engineering, Kyushu University, Fukuoka 819-0395, Japan
**Yabegawa Electric Industry Ltd., Oumuta 836-0844, Japan
***Department of Applied Chemistry, Faculty of Engineering, Shibaura Institute of Technology, Toyosu, Koto, Tokyo 135-8548, Japan

A rapid, sensitive immunoassay based on a surface plasmon resonance sensor in a flow system for the determination of alkylphenol polyethoxylate (APEO) is described. The method is based on an indirect competitive reaction between an anti-APEO antibody in the sample solution and APEO immobilized on a sensor chip and APEO in the same sample solution. A sensor chip was prepared by immobilizing an APEO-horseradish peroxidase (APEO-HRP) conjugate on the thin gold film of the sensor chip. The adsorption constants for the APEO-HRP conjugate on the sensor chip and the surface density of the APEO-HRP adsorbed on the sensor chip at the saturated state were estimated to be 4.7 × 10⁵ M⁻¹ and 5.0 × 10⁻¹⁴ mol/mm², respectively, using a Langmuir adsorption isotherm equation and results from the adsorption experiments. The affinity constants for the immunocomplexes of the anti-APEO antibody with the APEO conjugate on the sensor chip and for APEO in the sample solution were estimated to 2.0 × 10⁶ and 5.1 × 10⁶ M⁻¹, respectively. A typical sigmoid calibration curve for APEO was obtained in the concentration range from 1 ppb to 1000 ppb. The detection limit, defined as the concentration of APEO, at which 85% of the sensor signal was observed, was ca. 10 ppb. The assay was applied to the determination of APEO in tap water in conjunction with a solid phase extraction pretreatment; APEO levels of approximately 50 ppt were successfully determined.

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