Capillary electrophoresis and isoelectric focusing in peptide and protein analysis

Abstract

CZE and CIEF of proteins have preceded, and accompanied, the birth of proteomics. Although they might not be fully exploited in massive proteomic analyses (especially those projects aiming at a deep discovery of possibly the entire proteome of a cell or subcellular organelles or biological fluids), it still has interesting features and advantages, especially with samples of limited heterogeneity and in the field of purity checking for recombinant DNA proteins meant for human consumption. The purpose of this tutorial paper is to guide the reader through the history of the field, then through the main steps of the process, from sample preparation to analysis of proteins and peptides, while commenting on the constraints and caveats of the technique. The tutorial ends with an outlook on the future, which might be dominated by microchip electrophoresis, especially for ultrafast analyses of protein samples in a sieving mode, in presence of either sieving liquid polymers or firm gels polymerized within the microchannels. To this purpose, commercial instrumentation is already available on the market. This tutorial is part of the International Proteomics Tutorial Programme (IPTP 13).

Direct and Indirect Applications of Sub- and Supercritical Water in Food-Related Analysis

“…An important portion of bioanalytes (e.g., amino acids, peptides and proteins) are of amphoteric nature. There are several variants of isoelectric focusing that differ in the format and/or medium of the separation space-gel isoelectric focusing, preparative (free flow) isoelectric focusing, and capillary isoelectric focusing (CIEF) (Righetti 2006;Silvertand et al 2008;Koshel and Wirth 2012;Righetti et al 2013). The latter variant is the subject here.…”

Recent advances in the analysis of therapeutic proteins by capillary and microchip electrophoresis

“…CIEF separates proteins based on their isoelectric point (pI) and can be used to determine charge heterogeneity of biogenic products [49]. The assay is typically performed in a coated capillary to eliminate EOF.…”

Gold-nanoparticle extraction and reversed-electrode-polarity stacking mode combined to enhance capillary electrophoresis sensitivity for conjugated nucleosides and oligonucleotides containing thioether linkers

“…High-performance capillary electrophoresis (HPCE) has proved to be a rapid and simple technique for separating charged biomolecules and nanoparticles with very high resolution [11]. Its unique advantages, including relatively short analysis time and high separation efficiency with consumption of a minimal amount of sample and buffer solution, make HPCE a valuable technique for the determination of biomolecules [12], especially when a small amount of sample is available for the analysis. Micellar electrokinetic chromatography (MEKC) methods with sodium dodecyl sulfate (SDS) and borate phosphate buffers have been used for the determination of nucleosides by many researchers [13][14][15].…”