Monoclonal antibodies (mAbs) have been successfully developed for the treatment of a number of serious diseases. The FDA has approved more than 20 mAbs, and an additional 150 mAbs are currently in preclinical or clinical trials, or awaiting FDA approval. In the last decade, mAbs have become the largest part of the growing biologics drug market and have transformed the biotechnology and biopharmaceutical industries by growing exponentially into annual multibillion dollar sales. One of the key challenges in the development of mAbs is the comprehensive analytical characterization required to demonstrate consistent product quality.
mAbs generally exhibit charge heterogeneity from C-terminal processing of lysine residues, deamidation, glycation, N-terminal pyroglutamate formation, and other modifications. Therefore, the manufacturing and testing procedures for mAbs involve the monitoring of impurities resulting from these modifications.
Weak cation-exchange chromatography is well suited for such applications, and over the years Dionex’ ProPac® WCX-10 columns, packed with nonporous polymeric particles with carboxylic acid functional groups, have been considered the gold standard for high-resolution analysis and characterization of acidic and basic variants using inert HPLC platforms.
In this article, we describe the MAbPac™ SCX-10 column with a new stationary phase with strong cation-exchange functionality, specifically developed for the characterization of heterogeneity of mAbs. This column complements the existing ProPac WCX-10 column, providing high resolution and orthogonal selectivity for mAb separations. We demonstrate selectivity differences in heterogeneity characterization of mAbs using both the ProPac WCX-10 and the MAbPac SCX-10 columns, emphasizing the application of these columns.
The MAbPac SCX-10 column is based on a 10 µm nonporous, highly cross-linked styrenic polymeric media with a uniform hydrophilic coating. Strong cation-exchange functionality is introduced by grafted polymeric chains with sulfonic acid groups. The ATRP grafting technique is used to control the degree of polymerization and graft density of these polymeric chains. As a result, this stationary phase exhibits stability over a wide pH range with high selectivity and minimal band spreading.