Analysis of Monoclonal Antibody Charge Variants by Capillary Zone Electrophoresis

Charge heterogeneity analysis is important in the characterisation of monoclonal antibodies because it provides important information about product quality and stability. Heterogeneity can be caused by such molecular adaptions as C-terminal lysine modification, deamidation, and post translational modification. One method for separating charge variants is capillary isoelectric focusing (cIEF) which provides information regarding isoelectric point variation for related molecular isoforms. cIEF requires electrophoretic separation in a coated capillary which helps suppress electro-osmotic flow (EOF) and prevents surface protein adsorption but can result in a fairly long cycle time and complex sample preparation.
We demonstrate here that by using a simple separation buffer system and a bare fused silica capillary, it is possible to obtain a highly resolved, reproducible separation of a representative monoclonal antibody in less than 12 minutes. We also demonstrate that while high resolution can be achieved using a short effective length (20cm), the fine structure of the monoclonal antibody used in this study can be revealed by increasing the effective length to 40cm.