Rho Hoon Suk, Hanke Alexander Thomas, Ottens Marcel, Gardeniers Han J G E
Mesoscale Chemical Systems Group, MESA+ Institute for Nanotechnology, University of Twente, Enschede, The Netherlands.
BioProcess Engineering group, Department of Biotechnology, Faculty of Applied Sciences, Delft University of Technology, Delft, The Netherlands.
Electrophoresis. 2018 Apr;39(8):1031-1039. doi: 10.1002/elps.201700341. Epub 2018 Feb 5.
A microfluidic device for pH gradient chromatofocusing is presented, which performs creation of a micro-column, pH gradient generation, and fraction collection in a single device. Using a sieve micro-valve, anion exchange particles were packed into a microchannel in order to realize a solid-phase absorption column. To fractionate proteins according to their isoelectric points, elution buffer solutions with a stepwise pH gradient were prepared in 16 parallel mixing reactors and flowed through the micro-column, wherein a protein mixture was previously loaded. The volume of the column is only 20 nL, hence it allows extremely low sample consumption and fast analysis compared with a conventional system. We demonstrated separation of two proteins, albumin-fluorescein isothiocyanate conjugate (FITC-BSA) and R-Phycoerythrin (R-PE), by using a microcolumn of commercial charged polymeric particles (Source 15Q). The microfluidic device can be used as a rapid diagnostic tool to analyse crude mixtures of proteins or nucleic acids and determine adsorption/desorption characteristics of various biochemical products, which can be helpful for scientific fundamental understanding as well as instrumental in various industrial applications, especially in early stage screening and process development.
本文介绍了一种用于pH梯度色谱聚焦的微流控装置，该装置可在单个设备中完成微柱创建、pH梯度生成和馏分收集。使用筛网微阀将阴离子交换颗粒填充到微通道中，以实现固相吸收柱。为了根据蛋白质的等电点对其进行分离，在16个平行混合反应器中制备了具有逐步pH梯度的洗脱缓冲溶液，并使其流过预先加载了蛋白质混合物的微柱。该柱的体积仅为20 nL，因此与传统系统相比，它具有极低的样品消耗和快速分析的特点。我们使用商业带电聚合物颗粒（Source 15Q）的微柱展示了两种蛋白质，即异硫氰酸荧光素标记的白蛋白（FITC-BSA）和藻红蛋白（R-PE）的分离。该微流控装置可用作快速诊断工具，用于分析蛋白质或核酸的粗混合物，并确定各种生化产物的吸附/解吸特性，这有助于科学的基础理解，并在各种工业应用中发挥作用，特别是在早期筛选和工艺开发中。
