Affinity Interactions in Capillary Separations

密歇根大学的Robert T. Kennedy教授和他的团队在化学系分析和表面化学项目的支持下，构思并实施了基于微型化电泳的测量蛋白质和蛋白质反应的新方法。工作包括开发一种高通量微流控“芯片”，它可以在载油流中以纳升体积的液滴收集水样，然后以每秒2-5的异常快的速度通过电泳分离液滴的内容物。作为一个模型系统，该小组正在筛选能够影响细胞信号传导中某些蛋白质-蛋白质相互作用的化合物。第二种方法是微尺度的Western blot，通过在凝胶板中筛分电泳分离蛋白质，转移到印迹膜上，然后通过与抗体的相互作用在膜上进行检测。基于微流体工具的微型、低成本和高度自动化的Western blot方法的潜在效用将通过测量胚胎分泌的蛋白质来证明，以检查它们在胚胎发育中的作用。这种新一代工具的发展预计将对生物技术、化学基因组学、药物发现、临床诊断和基础生物化学等领域产生影响。这些工具将有助于根据干扰生物化学相互作用的能力筛选化合物库，从而发现生物网络的化学探针或新药。开发的Western blot方法将使微尺度样品的高通量成为可能。如果成本足够低，该方法也可以应用于低资源环境。这项工作为研究生和博士后提供了跨学科的培训机会，使他们能够在各个领域做出贡献，特别是在生物技术和生物分析化学领域。预计参与该项目的学生将在工业界或学术界担任职务，从而为国家科学基础设施做出贡献。

Professor Robert T. Kennedy and his group at the University of Michigan are supported by the Analytical and Surface Chemistry Program in the Division of Chemistry to conceive and implement new methods of measuring proteins and protein reactions based on miniaturized forms of electrophoresis. Work includes development of a high-throughput microfluidic "chip" that can collect aqueous samples as nanoliter volume droplets in a carrier stream of oil and then separate the contents of the droplets by electrophoresis at unusually fast rates of 2-5 per second. As a model system, the group is screening for compounds that can affect certain protein-protein interactions involved in cell signaling. A second thrust involves a microscale Western blot by which proteins are separated by sieving electrophoresis in slabs of gel, transferred to a blotting membrane, and then detected on the membrane by interaction with an antibody. Potential utility of the resulting miniaturized, low-cost and highly automated Western blot method based on microfluidic tools will be demonstrated by measuring proteins secreted from embryos in order to examine their role in embryo development. Development of this new generation of tools is expected to have impact in fields as diverse as biotechnology, chemical genomics, drug discovery, clinical diagnostics and fundamental biochemistry. The tools will be useful to screen libraries of compounds based on their ability to perturb biochemical interactions, enabling discovery of chemical probes of biological networks or novel drugs. The Western blot method developed will enable high-throughput on microscale samples. If sufficiently low cost, the method may also have application in low-resource environments. The work provides interdisciplinary training opportunities for graduate and post-doctoral students, preparing them to make contributions in a variety of fields, but especially in the areas of biotechnology and bioanalytical chemistry. It is anticipated that students working on this project will take positions in industry or academia thus contributing to national science infrastructure.