基于表面等离子体共振的小分子竞争与信号增强免疫反应痕量检测

Surface plasmon resonance-based trace detection of small molecules by competitive and signal enhancement immunoreaction.

作者信息

Aizawa Hidenobu, Tozuka Mitsuhiro, Kurosawa Shigeru, Kobayashi Koichi, Reddy Subrayal M, Higuchi Masahiro

机构信息

National Institute of Advanced Industrial Science & Technology, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8565, Japan.

出版信息

Anal Chim Acta. 2007 May 22;591(2):191-4. doi: 10.1016/j.aca.2007.03.074. Epub 2007 Apr 7.

DOI:10.1016/j.aca.2007.03.074

PMID:17481407

Abstract

A surface plasmon resonance (SPR)-immunosensor for detection of the low molecular weight compound 2,4-dinitorophenol (DNP) at ultra-low concentration has been developed. The sensor strategy is based on a competitive immunoreaction between DNP and a DNP-protein conjugate, namely DNP-bovine serum albumin conjugate (DNP-BSA). Anti-DNP monoclonal antibody was immobilized on a gold thin-film coated SPR-sensor chip by means of a chemical coupling process. DNP-BSA, on contact with the anti-DNP antibody immobilized SPR-immunosensor chip causes an increase in the resonance angle of the sensor chip. The optimum concentration of immobilized antibody on the SPR-sensor chip is 100 microg mL(-1). The SPR-immunosensor response for free DNP determination using the competitive immunoreaction had a response time of ca. 15 min. Using this method, DNP could be determined in the concentration range 1 ppt to 1 ppb. The SPR signal for ppt levels of DNP was enhanced by a factor of three by subsequently treating immuno-bound DNP-BSA with a secondary anti-DNP antibody.

摘要

已开发出一种用于检测超低浓度低分子量化合物2,4-二硝基苯酚（DNP）的表面等离子体共振（SPR）免疫传感器。该传感器策略基于DNP与DNP-蛋白质偶联物（即DNP-牛血清白蛋白偶联物，DNP-BSA）之间的竞争性免疫反应。通过化学偶联过程将抗DNP单克隆抗体固定在涂有金薄膜的SPR传感器芯片上。DNP-BSA与固定有抗DNP抗体的SPR免疫传感器芯片接触会导致传感器芯片的共振角增加。SPR传感器芯片上固定抗体的最佳浓度为100μg mL(-1)。使用竞争性免疫反应进行游离DNP测定的SPR免疫传感器响应时间约为15分钟。采用该方法，DNP的测定浓度范围为1ppt至1ppb。通过随后用抗DNP二抗处理免疫结合的DNP-BSA，使ppt水平的DNP的SPR信号增强了三倍。