Development of a capillary gel electrophoresis method for monitoring disulfide isomer heterogeneity in IgG2 antibodies

Abstract

A CGE method for monitoring the disulfide isomer distribution characteristic of IgG2 MAbs is presented. Disulfide heterogeneity of MAbs has been studied using various chromatographic and electrophoretic methods. Although CGE operates using a different selectivity mechanism from that of sorption chromatographic techniques, similar trends are present in the data, which allow the CGE method to be used as a complementary method for studying disulfide isomer distribution. This article focuses on the optimization of a capillary-based gel electrophoresis method that can be used to support antibody development including bioprocess optimization, antibody characterization, release, and formulation stability assessment.

Ionic liquids as buffer additives in ionic liquid-polyacrylamide gel electrophoresis separation of mixtures of low and high molecular weight proteins

“…27,65,66 In contrast, in IL-PAGE, separation is based on the charge on the proteins and their molecular masses. 27,65,66 In contrast, in IL-PAGE, separation is based on the charge on the proteins and their molecular masses.…”

Protein separation by capillary gel electrophoresis: A review

“…Zhang et al [48] optimized a similar method to analyze mAb1 drug substance under both reduced and non-reduced conditions. Lancher et al [127] established a generic method for monitoring disulfide isomer heterogeneity in IgG2 antibodies, and applied this method for purity analysis of reduced and non-reduced IgG2 mAbs [128]. Rustandi et al [129] reported a wide range of applications of CGE for mAb product development, including purity analyses for product release, product-related impurities during process and cell-culture development, and product stability evaluation.…”

Ionic liquid‐assisted SDS‐PAGE to improve human serum protein separation

“…BME or DTT in SDSsample buffer destroyed the disulfide bond between the amine acids in protein and eliminated the protein's original form difference. So all proteins would have roughly the same mass-to-charge ratio, and existed in the form of SDS-protein complex [15,38,40]. The formation of ILs-PAG was based on the reaction of AAm and Bis, cation in ILs could be embedded into the gel, and the positive charge and hydrophobic alkyl chain in ILs cation could provide some electrostatic and hydrophobic interactions with the passing SDS-protein complex, which played an important role in changing their electrophoretic mobility.…”