Column coupling isotachophoresis–capillary electrophoresis with mass spectrometric detection: Characterization and optimization of microfluidic interfaces

Two-dimensional electrophoretic separations are one of the most promising tools for the continuously growing needs of different bioanalytical fields such as proteomics and metabolomics. In this work we present the design and the implementation of a two-dimensional electrophoretic separation coupled to mass spectrometry. We started our work studying the sample transfer characteristics of different microfluidic interfaces compatible with capillary coupling for two-dimensional electrophoretic separations. These junctions are aimed at method decoupling and sample transfer in a modular two-dimensional electrophoretic separation system. In order to perform the characterization of the interfaces, we carried out capillary electrophoresis experiments and numerical simulations using three cationic compounds under different flow conditions. The comparison of the experimental and simulation results enables us to clearly define the desirable characteristics of interfaces in order to achieve method orthogonality with lossless sample transfer in a two-dimensional separation system. Finally, we present a glass microfluidic chip as interface for the implementation of a novel hybrid modular system for performing two-dimensional electrophoretic separations involving isotachophoresis and capillary electrophoresis. In this setup we include mass spectrometric and contactless capacitively coupled conductivity detection to monitor the separation process. We demonstrate the ability of the setup to be used as a flexible analysis tool by performing preconcentration, separation, detection and identification of four different human angiotensin peptides.

•Characterization of two different interfaces for 2D electrophoretic separations.•Interfaces were modeled using a novel extended 1D FEM numerical method.•Microfluidic chips shown excellent performance for sample transfer between columns.•Novel hybrid capillary-microchip concept is presented and tested for ITP–CE separations.•Setup was coupled to TOF-MS for separation and identification of human peptides at lower concentrations compared to previous studies.