Optimization of Immobilized Bacterial Disaccharides for Surface Plasmon Resonance Imaging Measurements of Antibody Binding
Optimization of Immobilized Bacterial Disaccharides for Surface Plasmon Resonance Imaging Measurements of Antibody Binding
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DOI：
10.1021/la8026489
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发表时间：
2008-12-16
期刊：
影响因子：
3.9
通讯作者：
McDermott, Mark T.
中科院分区：
文献类型：
--
作者：
Grant, Chris F.;Kanda, Vishal;McDermott, Mark T.
蛋白质和固定化碳水化合物之间的相互作用对细胞信号传导和免疫反应等生物事件至关重要。用碳水化合物修饰表面以创建传感平台为在实验室环境中研究这些相互作用提供了一条途径。在这项工作中，一个家庭的结构相关的沙门氏菌二糖表位固定在薄的金膜上的阵列格式探测抗体结合表面等离子体共振（SPR）成像。二糖用烷基硫醇接头修饰，以便于固定到金上。固定化的修饰的二糖的立体化学中的微小差异显示出极大地影响单克隆抗体的结合。具体而言，仅观察到相对于泰维糖或帕拉糖类似物与固定的阿贝可糖双脱氧己糖的结合。然而，结合抗体的量和相对强度取决于二糖部分在表位和非结合稀释剂分子的混合单层中的分布。我们用各种技术彻底表征混合单层，以了解用于抗体捕获的二糖的最佳密度和分布。这项工作加强了在界面处控制配体密度以优化基于表面的生物测定的重要性。
The interactions between proteins and immobilized carbohydrates are crucial to biological events such as cell signaling and immune response. The modification of surfaces with carbohydrates to create sensing platforms provides a pathway to study these interactions in a laboratory setting. In this work, a family of structurally related Salmonella disaccharide epitopes is immobilized on thin gold films in an array format to probe antibody binding with surface plasmon resonance (SPR) imaging. The disaccharides are modified with an alkyl thiol linker for facile immobilization to gold. Small differences in the stereochemistry of the immobilized, modified disaccharides are shown to greatly influence the binding of a monoclonal antibody. Specifically, binding is only observed to an immobilized abequose dideoxyhexose relative to a tyvelose or a paratose analogue. However, both the amount and relative strength of bound antibody depends on the distribution of disaccharide moieties in a mixed monolayer of the epitope and a nonbinding diluent molecule. We thoroughly characterize the mixed monolayers with a variety of techniques to understand the optimal density and distribution of the disaccharide for antibody capture. This work reinforces the importance of controlling the density of ligands at the interface for optimized surface based bioassays.