High-throughput Carbonyl Content Method of Therapeutic mAb using size-exclusion chromatography with ultraviolet and fluorescence detection
2019
Yi, Yunyu | Zang, Li
Monoclonal antibody (mAb), one of the major types of therapeutic proteins in the pharmaceutical industry, is predominantly manufactured using mammalian cell culture [1]. Oxidative stress, potentially present during cell culture process, may increase the protein carbonyl content in the mAb product, which was reported to positively correlate with aggregate burst rate during storage [2]. In order to monitor carbonyl content during mAb process development, we developed a high-throughput screening method for therapeutic mAbs using size-exclusion chromatography followed by ultraviolet and fluorescence detection (SEC-UV/FL), optimized from a fluorescein thiosemicarbazide (FTC) semi-microplate method. The method demonstrated a good correlation with conventional ELISA assay and FTC-based fluorometric semi-microplate method with improved throughput and precision. The method was successfully applied in three case studies to improve our understanding of mAb carbonylation, including the impact of metal-catalyzed oxidation on an IgG4 mAb, comparison of carbonyl content between several mAbs expressed by CHO cell culture with human serum antibody pool, as well as the surface charge property of carbonylated mAb assessed by ion-exchange chromatography.
Mostrar más [+] Menos [-]Palabras clave de AGROVOC
Información bibliográfica
Este registro bibliográfico ha sido proporcionado por National Agricultural Library