Micro and Nanofluidic Methods for Studying Supramolecular Assembly

该项目的重点是发展用于研究超分子蛋白质组装的微流体和纳米流体方法。我们的目标是确定在一个复杂的混合物中的蛋白质和蛋白质复合物，通过捕获它们，使用电泳，静电和流体动力学的力量，在nanojunctions上的微流控设备制造使用软光刻技术。这种方法将允许选择性富集感兴趣的物种，从而使我们即使在稀释的分析物溶液中也能达到非常高的灵敏度，从而允许鉴定错误折叠的蛋白质和不正确组装的蛋白质复合物的少数群体。作为该项目的一部分，我们将开发在芯片上有效集成纳米和微流体元件的方法。这一目标将通过使用双光子光刻来实现，以克服通过非线性光学器件的衍射极限，并在用于常规微加工的相同光致抗蚀剂中定义纳米级特征。然后，光刻图案化的衬底将用于在软光刻工艺中压印聚合物，以产生可扩展的途径来制造集成纳米和微米尺度特征的器件。我们将通过深紫外荧光检测蛋白质，这将使我们能够与完全未标记的物种合作。这些结果将开辟了在其天然环境中以高分辨率定量表征蛋白质自组装的道路。

This project focuses on the development of micro and nanofluidic methods for the study of supramolecular protein assembly. We will aim to identify within a complex mixture proteins and protein complexes by trapping them, using electrophoretic, electrostatic and hydrodynamic forces, at nanojunctions on microfluidic devices fabricated using soft lithographic techniques. This approach will allow the selective enriching of species of interest and will thus allow us to reach very high sensitivities even with dilute analyte solutions, allowing minority populations of misfolded proteins and incorrectly assembled protein complexes to be identified. As part of the project we will develop approaches for effective integration of nano and microfluidic elements on chip. This objective will be achieved through the use of two-photon lithography to overcome the diffraction limit through non-linear optics and to define nanoscale features in the same photoresist used for conventional microfabrication. The lithographically patterned substrate will then be used to imprint polymers in a soft lithography process to yield a scalable avenue to fabricate devices integrating nano and micro scale features. We will detect proteins through deep UV fluorescence, which will allow us to work with fully unlabelled species. These results will open up the path towards quantitative charaterisation of protein self-assembly at high resolution in their native environment.