Ultra-High-Pressure LC (UHPLC) for Therapeutic Protein Characterization

摘要

HPLC is the most widely used technique in the biopharmaceutical industry for characterizing protein heterogeneity. The introduction of UHPLC provides an opportunity to increase speed and resolution for the chromatographic separation of proteins. This paper describes three UHPLC applications for protein characterization. The first application employs RP-HPLC to separate the free thiol variants of multiple recombinant monoclonal antibodies (mAb) in 2.5 minutes. The second application is an improved peptide mapping method using UHPLC to monitor the oxidation of different methionines for a mAb in about 69 minutes, instead of 216 minutes using a regular HPLC. Lastly, UHPLC-SEC method has been used for the separation of an E-coli derived Fab and its dimer form in four minutes. These high throughput and high efficiency UHPLC methods demonstrate the potential of UHPLC for protein characterization.