Brod E, S Ben-Yosef V, Bandhakavi S, Sivan U
Department of Physics and the Russell Berrie Nanotechnology Institute, Technion-Israel Institute of Technology, Haifa 3200003, Israel.
Bio-Rad Laboratories, Life Science Group, Hercules, CA 94547, United States.
J Chromatogr A. 2016 Jan 29;1431:166-175. doi: 10.1016/j.chroma.2015.12.070. Epub 2015 Dec 30.
A new method for generating complex, dynamic pH profiles in an ampholyte-free separation channel is presented together with the theory behind its operation. The pH is modulated by an array of proton and hydroxide ion injectors placed along the separation channel. The ions generated in-situ by electrically driven water splitting across a bipolar membrane are injected to the channel in the presence of a longitudinal electric field, leading to the formation of a multi-step pH profile. Real-time control over the pH profile along the channel facilitates new dynamic separation strategies as well as steering and harvesting of focused molecules, which are both impossible with conventional separation methods. These freedoms are particularly attractive for Lab-on-a-Chip applications. The pH step-like profile alleviates one of the main hurdles of conventional isoelectric separation methods, namely, the slowing down of focused molecules as they approach their focusing spot. As a result, separation is completed within minutes for both peptides and proteins, even with low applied electric fields. We demonstrate protein and peptide separation within minutes, and resolution of ΔpI=0.2. Novel separation strategies based on spatio-temporal pH control are demonstrated as well.
本文介绍了一种在无两性电解质分离通道中生成复杂动态pH分布的新方法及其运行背后的理论。pH值由沿分离通道排列的一系列质子和氢氧根离子注入器调节。通过双极膜上的电驱动水分解原位产生的离子在纵向电场存在的情况下被注入到通道中，从而形成多步pH分布。对沿通道的pH分布进行实时控制有助于实现新的动态分离策略以及聚焦分子的引导和收集，而这些对于传统分离方法来说都是不可能的。这些优势对于芯片实验室应用尤其具有吸引力。pH阶梯状分布缓解了传统等电分离方法的一个主要障碍，即聚焦分子在接近其聚焦点时速度减慢。因此，即使在低电场应用下，肽和蛋白质的分离也能在几分钟内完成。我们展示了几分钟内蛋白质和肽的分离以及ΔpI = 0.2的分辨率。还展示了基于时空pH控制的新型分离策略。
