摘要
Scaffolded DNA origami, a versatile method to construct high yield self- assembled DNA nanostructures, has been investigated to develop water-soluble nanoarrays for label free RNA detection, drug delivery, molecular positioning and recognition, and spatially ordered catalysis of single molecule chemical reactions. Its attributes that facilitate these applications suggest DNA origami as a candidate platform for intracellular targeting. After the interaction with targeted proteins in cell lysate, it is critical to separate and concentrate DNA origami nanoarrays from the crude cell lysate for further analysis. The recent development of microchip isotachophoresis （ITP） provides an alternative robust sample preconcentration and electrophoretic separation method. In this study, we present online ITP for stacking, separation and identification of aptamer-functionalized DNA origami and its thrombin complex in a simple cross-channel fused silica microfluidic chip. In particular, the method achieved separation of a binding complex in less than 5 min and 150-fold signal enhancement. We successfully separated and analyzed the thrombin bound origami-aptamer spiked into cell lysate using on-chip ITP. Our results demonstrate that origami/thrombin nanostructures can be effectively separated from cell lysate using this method and that the structural integrity of the concentrated binding complex is maintained as confirmed by atomic force microscopy （AFM）. An ITP-based separation module can be easily coupled to other microchip pre- and post-processing steps to provide an integrated proteomics analysis platform for diagnostic applications.
Scaffolded DNA origami，构造高收益 selfassembled DNA nanostructures 的一个万用的方法，被调查了为标签开发水溶性的 nanoarrays 免费 RNA 察觉，药交货，分子的放和识别，和单个分子化学药品反应的空间地订的催化作用。它便于这些应用程序的属性为细胞内部的指向作为一个候选人平台建议 DNA origami。在和在房间 lysate 的指向的蛋白质的相互作用以后，为进一步的分析把 DNA origami nanoarrays 与粗略的房间 lysate 分开并且专注是批评的。微芯片 isotachophoresis (ITP ) 的最近的发展提供一个其他的柔韧的样品 preconcentration 和 electrophoretic 分离方法。在这研究，我们在简单跨隧道的熔化硅石 microfluidic 的 aptamer-functionalized DNA origami 和它的凝血酵素建筑群的为叠的现在的联机 ITP，分离和鉴定削。特别地，方法在不到 5 min 和 150 褶层信号改进完成了有约束力的建筑群的分离。我们成功地分开了并且分析用在薄片上 ITP 刺进房间 lysate 的凝血酵素界限 origami-aptamer。我们的结果证明 origami/thrombin nanostructures 能有效地用这个方法与房间 lysate 被分开并且集中的有约束力的建筑群的结构的正直是由原子力量显微镜学(AFM ) 证实了被维持。一个基于 ITP 的分离模块能容易被联合到为诊断应用程序提供一个综合 proteomics 分析平台的另外的微芯片预处理和 processing 以后步骤。
