New Materials for High Efficiency Protein Separations

我们的目标是研究一类新的材料，有潜力提供一个显着的 与聚合物凝胶相比，蛋白质电泳的改进。今天的凝胶由于凝胶中的不均匀性而产生不可再现位置的扭曲的拖尾区，并且它们由于与有机溶剂的差的相容性而妨碍疏水蛋白的分离。拟议的工作是研究由二氧化硅纳米球有序阵列组成的材料，这些二氧化硅纳米球具有束缚在其表面的纳米级聚合物链。间隙体积充当筛分介质，并且聚合物链抵抗蛋白质吸附。由二氧化硅纳米球施加的结构定义承诺在蛋白质电泳中的高分离效率和再现性。拟议的工作包括三个部分。首先，要研究二氧化硅纳米球的结晶膜在板上和微流控芯片的通道中的受控沉积。聚合物涂层和纳米球的化学组成将有所不同，以了解如何最大限度地提高蛋白质的分离效率和聚合物和基底的碱稳定性。其次，将研究蛋白质通过这些介孔介质的筛分运输。分子量标准品和荧光标记蛋白的迁移速率和区域分布将被表征，以开发通过 有序中。将通过荧光相关光谱法探测材料的吸附特性，以确定蛋白质的移动的和非移动的部分，以及任何解吸过程的速率常数。第三，将研究等电聚焦以了解聚合物官能团、载体溶剂和固定物的位置在促进蛋白质溶解中的作用。 荧光相关光谱法将探测移动的和非移动的部分。将研究二维电泳分离，主要是筛分和等电聚焦与凝胶进行比较，也筛分和孔径梯度，以及电泳电色谱。 拟议的工作解决了蛋白质通过结晶介孔介质传输的核心问题，为开发强大的新蛋白质分离奠定了基础。

The goal is to investigate a new class of materials that has to potential to provide a significant improvement in protein electrophoresis, compared to polymer gels. Today's gels give distorted, tailing zones of irreproducible position due to inhomogeneities in the gel, and they hamper the separation of hydrophobic proteins by their poor compatibility with organic solvents. The proposed work is to investigate materials comprised by ordered arrays of silica nanospheres having nanometer-scale polymer chains tethered to their surfaces. The interstitial volume serves as a sieving medium, and the polymer chains resist protein adsorption. The structural definition imposed by the silica nanospheres promises high separation efficiency and reproducibility in protein electrophoresis. The proposed work entails three parts. First, the controlled deposition of crystalline films of silica nanospheres onto plates and into channels of microfluidic chips is to be investigated. The chemical compositions of polymer coating and the nanosphere will be varied to learn how to maximize separation efficiency for proteins and base stability for the polymer and substrate. Second, the sieving transport of proteins through these mesoporous media will be studied. The migration rates and zone profiles for molecular weight standards and for fluorescence-labeled proteins will be characterized to develop a model for transport through the ordered medium. The adsorptive properties of the material will be probed by fluorescence correlation spectroscopy to determine the mobile and immobile fractions of proteins, and the rate constants for any desorption processes. Third, isoelectric focusing will be investigated to understand the roles of polymer functional group, carrier solvent, and position of the immobiline, in promoting protein solubility. Fluorescence correlation spectroscopy will probe mobile and immobile fraction. Two-dimensional electrophoretic separations will be investigated, primarily sieving and isoelectric focusing for comparison with gels, and also sieving and pore-size gradients, and electrochromatography with electrophoresis. The proposed work addresses the central issues in protein transport through crystalline mesoporous media to lay the groundwork for development of powerful new protein separations.