Protein variant separations by cation-exchange chromatography on tentacle-type polymeric stationary phases

We developed a set of prototype cation-exchange column packings that are ba sed on a hydrophilic coated, pellicular polymeric support with a grafted te ntacular surface chemistry that is highly suited to resolving closely relat ed protein variants. These column packings (1) afford minimal band spreadin g in conjunction with extremely high selectivity, (2) exhibit a very hydrop hilic character and (3) have moderate loading capacity. Cytochrome c varian ts (bovine, horse, rabbit) were baseline-separated, as was native ribonucle ase A and its two deamidation products, the Asp(67) and isoAsp(67) forms. H umanized monoclonal antibody variants differing in the presence of lysine a t the C terminus of the heavy chains were baseline-resolved. Finally, the s eparation of hemoglobin variants found in a sample containing elevated leve ls of glycated hemoglobin was also demonstrated. (C) 1998 Published by Else vier Science B.V. All rights reserved.