Improvement in protein separation in Barretts esophagus samples using two-dimensional capillary electrophoresis...

In two-dimensional capillary electrophoresis (2DCE) components are separated based on their size and hydrophobicity. A preliminary run separates analytes in the first capillary based on size (CSE). Following that, fractions are electrokinetically transferred across an interface into a second capillary, where components are further resolved according to hydrophobicity. In order to succeed in this analysis, two orthogonal methods should be selected for the different modes. The transfers from the first to the second capillary must be efficient in order to reduce tailing effects and lost of resolution. We report a new method to improve the resolution with our 2DCE instrumentation using CD doped buffers. When methyl beta cyclodextrin (mbetaCD) is added to the 2DCE interface buffer a stacking effect is described in the transfers from the first to the second dimension. In addition to that, changes in retention times are observed when proteins form complex with CD's helping in the separation. Protein fingerprints were obtained from BE homogenates using this method in presence of methyl beta cyclodextrin (mbetaCD). Within-day and between-day precision has been studied in order to establish the reproducibility of the methodology proposed.