Free-flow zone electrophoresis of peptides and proteins in PDMS microchip for narrow pI range sample prefractionation coupled with mass spectrometry.
Free-flow zone electrophoresis of peptides and proteins in PDMS microchip for narrow pI range sample prefractionation coupled with mass spectrometry.
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DOI：
10.1021/ac9025219
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发表时间：
2010-03-15
影响因子：
7.4
通讯作者：
Han, Jongyoon
中科院分区：
文献类型：
--
作者：
Song, Yong-Ak;Chan, Michael;Celio, Chris;Tannenbaum, Steven R.;Wishnok, John S.;Han, Jongyoon
在本文中，我们正在评估的战略排序肽/蛋白质的基础上的电荷质量，而不诉诸两性电解质和/或等电聚焦，使用一个单一的和两个步骤的自由流动区带电泳。我们开发了一种简单的制造方法，通过在玻璃基底上表面印刷疏水层来在PDMS芯片中创建用于自由流动区带电泳的盐桥。由于表面印刷的疏水层防止PDMS芯片和基底之间的等离子体结合，因此可以为自由流动区带电泳创建电结间隙。使用该装置，我们证明了在标准缓冲液系统中在给定pH下分离阳性和阴性肽和蛋白质，并使用LC/MS验证了分选结果。此外，我们通过芯片外滴定耦合两个分选步骤，并从样品混合物中分离出特定pI范围内的肽，其中pI范围仅由缓冲溶液的pH值设定。这种自由流动区带电泳分选装置，其简单的制造，和0.5 pH单位的分选分辨率，可能是一个高通量的样品分馏工具，靶向蛋白质组学使用LC/MS。
In this paper, we are evaluating the strategy of sorting peptides / proteins based on the charge to mass without resorting to ampholytes and / or isoelectric focusing, using a single- and two-step free-flow zone electrophoresis. We developed a simple fabrication method to create a salt bridge for free-flow zone electrophoresis in PDMS chips by surface printing a hydrophobic layer on a glass substrate. Since the surface-printed hydrophobic layer prevents plasma bonding between the PDMS chip and the substrate, an electrical junction gap can be created for free-flow zone electrophoresis. With this device, we demonstrated a separation of positive and negative peptides and proteins at a given pH in standard buffer systems, and validated the sorting result with LC/MS. Furthermore, we coupled two sorting steps via off-chip titration, and isolated peptides within specific pI ranges from sample mixtures, where the pI range was simply set by the pH values of the buffer solutions. This free-flow zone electrophoresis sorting device, with its simplicity of fabrication, and a sorting resolution of 0.5 pH unit, can potentially be a high-throughput sample fractionation tool for targeted proteomics using LC/MS.
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影响因子：
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