chapter  1
46 Pages

Mixed Beds

Chromatographic separations for proteins have been essentially operated around interactions between the solid-phase sorbents and protein analytes. Thanks to polypeptide properties, fundamental interactions have been played around ion exchange effects, hydrophobic associations, metal chelation, and dipole-dipole, singularly or sequentially. They are all classiˆed as single-mode sorbents since normally they display only

1.1 Introduction ......................................................................................................1 1.2 Crossing over Single Molecular Interactions: From Mixed-Mode

to Mixed-Bed Interaction .................................................................................4 1.3 Enhancement of Very Low Abundance Proteins ........................................... 10 1.4 Separation of Protein Categories .................................................................... 16

1.4.1 Removal of High-Abundance Proteins ............................................... 17 1.4.2 Concomitant Separation of Protein Groups with Similar

Types of Molecular Interactions ......................................................... 18 1.5 Mixed Beds as a Source of Selective Ligand Identiˆcation for

Afˆnity Chromatography ...............................................................................20 1.6 Mixed Beds for Isoelectric Group Separation ................................................25 1.7 “Blind” Protein Puriˆcation Processes with Mixed Beds ..............................28 1.8 The Place of Mixed-Mode Chromatography within a Protein Separation

Scheme ............................................................................................................ 31 1.8.1 Polishing Aspect or Removal of Impurity Traces from

Puriˆed Biologicals ............................................................................ 32 1.8.2 Selection of Media for Mixed Beds .................................................... 35 1.8.3 Streamlining Single-Bed with Mixed-Bed Chromatography .............36