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Silica colloidal crystals as a gel media replacement.

机译：硅胶晶体作为凝胶介质的替代物。

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摘要

Rigid pore networks formed using nonporous silica particles were used as an alternative separation medium for gel based electrophoretic techniques to enable high-throughput protein separations by reducing separation lengths. The miniaturization of isoelectric focusing to a length as short as 5 mm, with higher resolution than a 100-mm IPG strip, is shown for microchannels packed with chemically modified silica particles. Isoelectric focusing of prostate specific antigen revealed more charge variants in a 10-mm channel than a commercial gel strip with a 100-mm long immobilized pH gradient, with each having the same electric field of 800 V·cm-1 and pH gradient of 3--10. Shortening the channel to 5 mm gave significant drift and compression of the pH gradient. Despite this, comparable resolution for the 5- and 10-mm channels was obtainable in under 10 minutes for the charge variants of prostate specific antigen when a shallower pH gradient of 6 -- 8 and a higher field of 1600 V·cm-1 were used for the 5 mm channel length.;Size based separations of proteins in silica colloidal crystals in separation lengths of less than 5 mm is characterized for particle diameters of nominally 350 and 500 nm. A model is developed that relates the reduced electrophoretic mobility to the experimentally measurable porosity. The model fits the data with no adjustable parameters for the case of silica colloidal crystals packed in capillaries, for which independent measurements of the pore radii were made from flow data. Band broadening rapidly increases as the pore radius approaches the protein radius, indicating that the main contribution to broadening is the spatial heterogeneity of the pore radius. These combined results support the notion that rigid pore networks formed using nonporous silica can be used as an alternative medium for gel based techniques, and facilitates the design of new separations that would benefit from miniaturization.
机译：使用无孔二氧化硅颗粒形成的刚性孔网络用作基于凝胶的电泳技术的替代分离介质，可通过减少分离长度来实现高通量蛋白质分离。对于填充有化学改性二氧化硅颗粒的微通道，等电聚焦的微型化使其长度短至5 mm，比100 mm IPG条具有更高的分辨率。前列腺特异性抗原的等电聚焦显示，与商品凝胶条相比，固定长度为100 mm的商业化凝胶条在10 mm通道中具有更多的电荷变异体，每个电场均具有800 V·cm-1的相同电场且pH梯度为3 --10。将通道缩短至5 mm，可明显漂移并压缩pH梯度。尽管如此，当较浅的pH梯度为6-8和较高的磁场为1600 V·cm-1时，对于前列腺特异抗原的电荷变异体，在10分钟内可以获得5和10 mm通道的可比分辨率。用于5 mm的通道长度。硅胶胶体晶体中分离长度小于5 mm的基于大小的蛋白质分离，其特征是标称直径为350和500 nm。建立了将降低的电泳迁移率与实验可测量的孔隙率相关的模型。对于填充在毛细管中的二氧化硅胶体晶体，该模型使用没有可调整参数的数据进行拟合，对于这种情况，可从流量数据中独立测量孔半径。随着孔半径接近蛋白质半径，条带扩展迅速增加，这表明对扩展的主要贡献是孔半径的空间异质性。这些综合结果支持了这样一种观念，即使用无孔二氧化硅形成的刚性孔网络可以用作基于凝胶的技术的替代介质，并有助于设计得益于小型化的新分离方法。

著录项

  • 作者

    Birdsall, Robert E.;

  • 作者单位

    Purdue University.;

  • 授予单位 Purdue University.;
  • 学科 Chemistry Inorganic.;Chemistry Analytical.
  • 学位 Ph.D.
  • 年度 2013
  • 页码 153 p.
  • 总页数 153
  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类
  • 关键词

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