High-performance capillary electrophoresis of SDS-protein complexes using UV-transparent polymer networks

Abstract

This paper demonstrates the use of UV-transparent replaceable polymer networks for the separation of SDS-protein complexes on the basis of molecular weight. First, the use of linear (i.e. non-cross-linked) polyacrylamide is shown to provide molecular separation of SDS-protein complexes. A study reveals such columns to yield significantly greater lifetime than cross-linked gels because of the flexibility of the noncovalently attached polymer chains. However, column lifetime was still found to be limited (approximately 20-40 injections), and detection at 214 nm was problematical because of the absorbance of polyacrylamide. UV-transparent polymer networks of dextran and PEG were substituted for polyacrylamide with successful molecular weight sieving of SDS-protein complexes at 214 nm. Due to their low to moderate viscosities, these networks could be routinely replaced leading to the possibility of hundreds of injections with a single column. Migration time reproducibilities of 0.5% RSD or less were found with replacement of the network. Using dextran, calibration plots of peak area vs concentration of standard protein were linear over the range of 0.5 microgram/mL up to at least 0.25 mg/mL. Furthermore, plasma samples could be directly utilized because of the strong solvating power of SDS. Rapid separation of protein mixtures are demonstrated with these UV-transparent polymer networks.