By Joseph Pate, Benjamin Kester, Claudia Berdugo, Yunsong Li, Victor Vinci
The 3M™ Emphaze™ AEX Hybrid Purifier is a novel purification tool that utilizes a Q-functional hydrogel and 0.2 μm membrane to reduce impurities in a biopharmaceutical downstream process. This technology has been reported to provide a 4-log reduction in DNA at the clarification stage, which can greatly improve the efficiency of the Protein A capture step1. While the advantages provided by the Emphaze™ AEX Hybrid Purifier are far-reaching, its Q-functionality presents a challenge for target proteins with a low isoelectric point (pI). Proteins under these conditions will usually bind to anion exchange ligands which can negatively impact the functionality of a clarification stage filter.
This study involved a Design of Experiment (DoE) based approach to investigate the effects of pH and conductivity on the turbidity, yield, and impurity clearance of the Emphaze™ AEX Hybrid Purifier and the efficiency of the subsequent Protein A capture step. The results were then used to determine optimum filtration conditions of one such low pI fusion protein, and whether elements of this technique can be extended to purification of a monoclonal antibody (mAb) with a higher pI. Filtration conditions for the low pI target protein were identified with a 4-fold greater yield over normal harvest filtration conditions and more than 99.6% removal of host cell protein (HCP) post Protein A chromatography. Application of these strategic elements to mAb filtration demonstrated an extended pH range at which post Protein A HCP reduction exceeds 99.9%, as well as up to a 28.7% increase in column binding capacity. For each molecule studied, filtration using the Emphaze™ AEX Hybrid Purifier resulted in greater HCP reduction and lower turbidity than unfiltered pools. This experimental plan provides a baseline to tailor specific application of the Emphaze™ AEX Hybrid Purifier to each individual protein or antibody.