Department of Chemical Systems and Engineering, Graduate School of Engineering, Kyushu University, Hakozaki, Higashi-ku, Fukuoka 812-8581, Japan.
Talanta. 2003 Jul 4;60(4):733-45. doi: 10.1016/S0039-9140(03)00139-5.
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 degrees 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.
一种基于表面等离子体共振（SPR）的生物传感器已被开发出来，用于使用间接竞争免疫测定法监测 2,4-二氯苯酚，这是一种已知的二恶英前体。SPR 传感器通过使用金结合多肽（GBP）和蛋白 G 将抗-（2,4-二氯苯酚）抗体固定在 SPR 传感器芯片的金薄层上来制造。通过将 2,4-二氯苯酚样品溶液注入包含 SPR 传感器的流动系统中，来测量基于抗原-抗体反应的 SPR 响应。在使用修饰后的传感器芯片的直接免疫测定系统中，当注入 25ppm 的 2,4-二氯苯酚溶液时，观察到的 SPR 角度变化小于 0.001 度，没有明显的变化。为了提高 SPR 传感器的灵敏度，使用间接竞争免疫测定法与 SPR 传感器系统结合使用，该系统使用与牛血清白蛋白（BSA）偶联的 2,4-二氯苯酚。在竞争测定中，将包含各种浓度（10-250ppb）的 2,4-二氯苯酚的 350ppm 2,4-二氯苯酚-BSA 缀合物溶液注入 SPR 传感器系统。与直接免疫测定相比，发现这种间接免疫测定的灵敏度非常高，估计检测限为 20ppb。通过比较使用 GBP 固定传感器芯片上的抗体的程序与另一种修饰程序（其中使用 BSA 代替 GBP 固定传感器芯片上的抗体），证实了使用 GBP 固定抗体可增强对 2,4-二氯苯酚的敏感性。从 SPR 响应中估计了 2,4-二氯苯酚及其缀合物与抗体的亲和力常数。
