In an effort to better understand the unique selectivities offered by multi-modal chromatographic systems, experiments were carried out with a variety of cation exchange and multi-modal ligands using model protein systems. Nuclear Magnetic Resonance (NMR) was employed to identify and map ligand interaction surfaces between the proteins and anionic and multi-modal ligands in solution. Key chemical features within the ligands that interact with the protein surfaces were identified and the number and locations of interaction sites on the protein were determined and evaluated. Site directed spin-labeling electron paramagnetic resonance (SDSL-EPR) was also performed using a number of cysteine mutants to determine the binding orientation of the wild type protein on both cation exchange and multi-modal resin surfaces. Finally, this information from both free solution and resin binding studies was employed to distinguish “affinity” type multi-modal interactions from multi-site non-specific interactions.