Site-directed antibody immobilization using a protein A-gold binding domain fusion protein for enhanced SPR immunosensing
Site-directed antibody immobilization using a protein A-gold binding domain fusion protein for enhanced SPR immunosensing
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DOI：
10.1039/c3an36498d
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发表时间：
2013-01-01
期刊：
影响因子：
4.2
通讯作者：
Lechuga, Laura M.
中科院分区：
文献类型：
--
作者：
de Juan-Franco, Elena;Caruz, Antonio;Lechuga, Laura M.
我们已经实施了一种新的策略，用于基于使用融合蛋白，蛋白A-金结合结构域（PAG）的抗体到金表面上的定向固定。PAG由与葡萄球菌蛋白A的免疫球蛋白结合结构域偶联的金结合肽（GBP）组成。该融合蛋白提供了抗体的容易且快速的定向固定，保留其天然结构，同时使抗原结合位点（Fab）自由暴露。使用这种固定化策略，我们已经证明了通过SPR的人生长激素的免疫传感的性能。检测限为90 ng mL（-1），芯片间变异性低于7%。这种方法与其他策略的直接固定抗体在金表面的比较表明，PAG的方法提供了增强的灵敏度。
We have implemented a novel strategy for the oriented immobilization of antibodies onto a gold surface based on the use of a fusion protein, the protein A-gold binding domain (PAG). PAG consists of a gold binding peptide (GBP) coupled to the immunoglobulin-binding domains of staphylococcal protein A. This fusion protein provides an easy and fast oriented immobilization of antibodies preserving its native structure, while leaving the antigen binding sites (Fab) freely exposed. Using this immobilization strategy, we have demonstrated the performance of the immunosensing of the human Growth Hormone by SPR. A limit of detection of 90 ng mL(-1) was obtained with an inter-chip variability lower than 7%. The comparison of this method with other strategies for the direct immobilization of antibodies over gold surfaces has showed the enhanced sensitivity provided by the PAG approach.