3D antibody immobilization on a planar matrix surface
3D antibody immobilization on a planar matrix surface
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3D 抗体固定在平面基质表面上
DOI：
10.1016/j.bios.2011.07.003
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发表时间：
2011-10-15
影响因子：
12.6
通讯作者：
Dai, Youzhi
中科院分区：
文献类型：
--
作者：
Feng, Bo;Huang, Shirong;Dai, Youzhi
每单位面积固定的活性捕获抗体的量对于开发有效的抗体芯片、生物传感器、免疫测定和其他分子识别技术至关重要。在这项研究中，我们提出了一种新的，但简单的方法为导向的抗体固定在高密度的基础上形成一个有序的，有组织的聚集的免疫球蛋白G（IgG）和葡萄球菌蛋白A（SPA）。在His-标签与Ni 2+螯合之后，抗体以三维（3D）方式固定在固体表面上，并且很好地暴露分析物受体位点，这导致分析信号的显著增强，检测灵敏度增加超过64倍。下拉测定证实IgG抗体只能通过SPA连接间接结合Ni-2+基质，其中His标签负责结合Ni 2+，同源结构域负责结合IgG Fc。只要抗体探针是哺乳动物IgG，本文提出的固定方法可能是一种有吸引力的策略，用于构建高性能抗体阵列和生物传感器。(C)2011爱思唯尔有限公司版权所有。
The amount of active capture antibodies immobilized per unit square is crucial to developing effective antibody chips, biosensors, immunoassays and other molecular recognition technologies. In this study, we present a novel yet simple method for oriented antibody immobilization at high density based on the formation of an orderly, organized aggregation of immunoglobulin G (IgG) and staphylococcal protein A (SPA). Following the chelation of His-tag with Ni2+, antibodies were immobilized on a solid surface in a three-dimensional (3D) manner and exposed the analyte receptor sites well, which resulted in a substantial enhancement of the analytical signal with more than 64-fold increase in detection sensitivity. Pull-down assays confirmed that IgG antibody can only bind to Ni-2+ matrix indirectly via a SPA linkage, where the His-tag is responsible for binding Ni2+ and homologous domains are responsible for binding IgG Fc. The immobilization approach proposed here may be an attractive strategy for the construction of high performance antibody arrays and biosensors as long as the antibody probe is of mammalian IgG. (C) 2011 Elsevier B.V. All rights reserved.