Self-assembled PEG monolayer based SPR immunosensor for label-free detection of insulin
Self-assembled PEG monolayer based SPR immunosensor for label-free detection of insulin
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DOI：
10.1016/j.bios.2006.06.012
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发表时间：
2007-02-15
影响因子：
12.6
通讯作者：
Miura, Norio
中科院分区：
文献类型：
--
作者：
Gobi, K. Vengatajalabathy;Iwasaka, Hiroyuki;Miura, Norio
基于表面等离子体共振（SPR）原理，研制了一种简单、快速的连续流动免疫传感器，其响应时间小于5 min，可检测低至1 ng ml（-1）（ppb）的胰岛素。将胰岛素共价结合到OEG-DCA的Au-硫醇盐单层上，用于激活传感器表面以进行免疫亲和相互作用。研究了一种在线竞争免疫传感原理用于检测胰岛素，其中在存在和不存在各种浓度的胰岛素的情况下，检测抗胰岛素抗体与传感器表面上的胰岛素的直接亲和结合。通过对抗体浓度、样品分析时间和流速等因素的优化，使抗胰岛素抗体与传感器表面的免疫反应达到理想的检测限和测定范围。该传感器对胰岛素的最低检测浓度为1 ng/ml，检测范围为1-300 ng/ml。所开发的基于OEG单层的传感器芯片对蛋白质的非特异性吸附表现出高抗性，并且已经证明了从胰岛素浸渍的血清样品中不间断地高灵敏度检测胰岛素。免疫反应循环后，简单地通过酸性缓冲液（甘氨酸）的短暂流动再生活性传感器表面。盐酸;发现相同的传感器芯片可重复使用超过25个循环，而原始传感器活性没有明显变化。(c)2006 Elsevier B. V.保留所有权利。
A simple and rapid continuous-flow immunosensor based on surface plasmon resonance (SPR) has been developed for detection of insulin as low as 1 ng ml(-1) (ppb) with a response time of less than 5 min. At first, a heterobifunctional oligo(ethyleneglycol)-dithiocarboxylic acid derivative (OEG-DCA) containing dithiol and carboxyl end groups was used to functionalize the thin Au-film of SPR chip. Insulin was covalently bound to the Au-thiolate monolayer of OEG-DCA for activating the sensor surface to immunoaffinity interactions. An on-line competitive immunosensing principle is examined for detection of insulin, in which the direct affinity binding of anti-insulin antibody to the insulin on sensor surface is examined in the presence and absence of various concentrations of insulin. Immunoreaction of anti-insulin antibody with the sensor surface was optimized with reference to antibody concentration, sample analysis time and flow-rate to provide the desired detection limit and determination range. With the immunosensor developed, the lowest detectable concentration of insulin is 1 ng ml(-1) and the determination range covers a wide concentration of 1-300 ng ml(-1). The developed OEG-monolayer based sensor chip exhibited high resistance to non-specific adsorption of proteins, and an uninterrupted highly sensitive detection of insulin from insulin-impregnated serum samples has been demonstrated. After an immunoreaction cycle, active sensor surface was regenerated simply by a brief flow of an acidic buffer (glycine. HCl; pH 2.0) for less than 1 min. A same sensor chip was found reusable for more than 25 cycles without an appreciable change in the original sensor activity. (c) 2006 Elsevier B.V. All rights reserved.