Table II. Comparison between co-elution separation strategies: SEC, IEX, HIC, BN-PAGE.

Separation technique	Stationary phase	Separation principle	Mobile phasea	Benefits/Drawbacks	Applications
SEC	Material with different pore sizes (silica, polymeric or cross-linked agarose)	Hydrodynamic size	Millimolar salt buffers at neutral pH	Less buffer requirements, isocratic/Modest resolution	Soluble complexes
IEX	Material with ionic groups (silica-based or polymeric) with SAX, WAX, WCX or mixed-bed (WAX & WCX) properties	Salt gradient	Increasing molar salt gradient (NaCl) at neutral pH	Higher resolution, more chemistries available/Higher salt concentrations required	Soluble complexes
HIC	Hydrophobic	Salt gradient	Decreasing molar salt gradient (e.g. (NH4)2SO4) at neutral pH	Higher salt concentrations required	Multiple orthogonal separations
BN-PAGE	Polyacrylamide gel	Electrophoretic mobility	Coomassie blue G, salt buffers	Not limited to soluble complexes/Less reproducible	Membrane protein complexes

^aSalts commonly used for buffers: NaCl, HEPES, KCl, MgCl₂, Tris, PBS, NaCH₃COO, NaN₃, (NH₄)₂SO₄. May contain additives like proteases inhibitors, dithiothreitol or glycerol.