Characterization of a capillary T-junction for multidimensional electrophoretic separations

KLER, PABLO A.; POSCH, TJORBEN N.; HUHN, CAROLIN

Ginebra

Congreso; 27th International Symposium on MicroScale Bioseparations and Analyses; 2012

Multidimensional electrophoretic separations consist of two or more independent separation steps arranged sequentially. Each step may use a different buffer composition and thus a different or even orthogonal separation mechanism. These provide a selective relative displacement between analytes based on different physiochemical properties of such analytes [1]. These electrophoretic separations comprise a number of electromigrative separation techniques as capillary electrophoresis (CE), isoelectric focusing (IEF), isotachophoresis (ITP) and micellar electrokinetic chromatography (MEKC), among others, that have been widely used independently or coupled [2-4], as is the case of IEF-sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), which has been used for several proteomics applications [5]. In order to obtain an efficient multidimensional separation two aspects have to be considered, the orthogonality (independence) of the mechanisms and a lossless sample transfer from one column to other. Some methods like ITP, IEF and CE are orthogonal when performed in separated columns [6]. Due to the different separation mechanisms involved, their coupling increases the need to develop an efficient technique/configuration for sample transfer between columns. In this work we present the modelling, simulation and validation of the hydrodynamic characteristics of a capillary T-junction. This junction is aimed to method decoupling and sample transfer in a two dimensional separation system involving ITP/CE or IEF/CE. Simulations were performed by Finite Element Methods [7]. CE experiments were carried out for three cationic compounds, with ammonium acetate as BGE (pH = 4.5), under different conditions of injection and flow. For injection we considered electrokinetic and pressure driven methods and the flow was driven by pure electroosmotic flow or combined with applied pressure. This characterization enables us to improve techniques and parameters for sample transfer on designing and implementing multidimensional electrophoretic separations.