Electrophoretic migration of proteins in semidilute polymer solutions

Abstract

We present a systematic study of the electrophoretic migration of 10-200 kDa protein fragments in dilute-polymer solutions using microfluidic chips. The electrophoretic mobility and dispersion of protein samples were measured in a series of monodisperse polydimethylacrylamide (PDMA) polymers of different molecular masses (243, 443, and 764 kDa, polydispersivity index <2) of varying concentration. The polymer solutions were characterized using rheometry. Prior to loading onto the microchip, the polymer solution was mixed with known concentrations of SDS (SDS) surfactant and a staining dye. SDS-denatured protein samples were electrokinetically injected, separated, and detected in the microchip using electric fields ranging from 100 to 300 V/cm. Our results show that the electrophoretic mobility of protein fragments decreases exponentially with the concentration c of the polymer solution. The mobility was found to decrease logarithmically with the molecular weight of the protein fragment. In addition, the mobility was found to be independent of the electric field in the separation channel. The dispersion is relatively independent of polymer concentration and it first increases with protein size and then decreases with a maximum at about 45 kDa. The resolution power of the device decreases with concentration of the PDMA solution but it is always better than 10% of the protein size. The protein migration does not seem to correspond to the Ogston or the reptation models. A semiempirical expression for mobility given by van Winkle fits the data very well.

Dynamic light‐scattering measurement of sieving polymer solutions for protein separation on SDS CE

“…The peak spacing increased and the mobility of proteins width decreased at x 5 10 nm or less. We approximated the average size of the proteins to be from 8 to 55 nm by taking the product of the average number of amino acids per protein and average length per amino acid [11]. Thus, the protein experiences the strong frictional force from polymer networks and the migration behavior of proteins changes as mesh size becomes smaller than the protein length.…”

“…Oliver and coworkers investigated the migration of protein through pDMA networks whose mesh size was estimated around 10-20 nm. Both the mobility of protein and the resolving power, that is, the smallest resolvable size difference, were improved with increasing concentration of pDMA [11]. In these studies, however, the mesh size of polymer network was calculated from the combination of polymer solution theory and viscosity measurement.…”

Rapid detection and quantification of specific proteins by immunodepletion and microfluidic separation

“…It is important to note that peak interference is a challenge in all the current methods available. More investigations are needed to improve resolution of the peaks by choosing different gel properties [16], or changing the script of the system. Release of only 19% of the FLAG protein from the beads shows that even though the target can be detected after elution, and further improvements are possible by optimizing the elution process, an accurate quantification is not possible by this method.…”

Electrokinetic motion of particles and cells in microchannels

“…In terms of important applications in single cell analysis, CE is an important tool for chemical cytometry and analysis of the intracellular components revealed by the resultant electropherograms (Stuart and Sweedler 2003;Desai and Armstrong 2003;Kremser et al 2004;Rodriguez and Armstrong 2004;Oliver et al 2008). Chen and Lillard (2001) demonstrated a high-throughput device for single-cell analysis by capillary electrophoresis (Fig.…”