Capillary sodium dodecyl sulfate gel electrophoresis of proteins: Introducing the three dimensional Ferguson method

One of the most extensively utilized rapid characterization, release and stability testing methods therapeutic proteins in biopharmaceutical field today is capillary SDS gel electrophoresis using borate cross-linked high molecular weight dextran. In spite its widespread use, however, composition dependent separation characteristics this very unique sieving matrix has not been investigated yet. Introduction three dimensional (3D) Ferguson plots, based on simultaneous variation dextran (D) (B) concentrations generating 16 different D/B ratio gels, allowed better understanding electromigration process SDS-protein complexes. As a result comprehensive study, non-linear 3D logarithmic mobility vs concentration surfaces were obtained. Both, protein standards new modality fusion etanercept resulted concave plots. The interplay between electroosmotic flow viscosity matrices played key role resulting migration time resolution. Selectivity values defined evaluated graph formats for regular de-N-glycosylated subunits etanercept, as well latter with 10 kDa internal standard to understand both dextran-borate complexation sized selectivities. KR plots retardation coefficients function weights used more precisely assess Mw samples obtain information about electromigrating