Development, validation, and implementation of capillary gel electrophoresis as a replacement for SDS‐PAGE for purity analysis of IgG2 mAbs

Abstract

Capillary gel electrophoresis (CGE) methods with UV detection were developed for reduced and non-reduced mAb analysis. These methods can be used to evaluate mAb purity, offering more reproducible quantitation compared with that of traditional SDS-PAGE methods. These CGE methods have been utilized as platform technology for bioprocess development, formulation development, mAb characterization, drug substance/drug product release testing as well as a required methodology for stability testing. We have found these CGE methods to be applicable across a platform of mAbs in preclinical and clinical development, with the majority of mAbs requiring no modification to the method conditions. This methodology has been ICH validated and transferred to several supporting organizations. The data presented herein describes the development of CGE methodology, platform application to mAb purity analysis, ICH validation, reliability metrics, and considerations on technology enhancement for improved performance and throughput.

Implementation of USP antibody standard for system suitability in capillary electrophoresis sodium dodecyl sulfate (CE-SDS) for release and stability methods

“…The disulfide bonds which are responsible for the tetrameric structure of monoclonal antibodies are disrupted (reducing) or protected (non-reducing) during heat denaturation by reducing or alkylating reagents respectively [15,16]. In the non-reducing condition, sensitivity of antibodies to partial reduction and fragmentation necessitate molecule dependent optimization of sample preparation [16][17][18].…”

Analysis of recombinant monoclonal antibodies by capillary zone electrophoresis

“…For example, peak tailing was observed when new‐coated capillaries were used, which has been interpreted to result from protein adsorption to the capillary surface . This phenomenon occurred in two of three newly installed capillaries, which might be an indication of the variability in capillary polyacrylamide‐based surface coating between batches.…”

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.…”