Size exclusion chromatography (SEC) is often plagued with electrostatic and hydrophobic secondary interactions. These affect the quality of the separation and ability to characterize and monitor important product-related impurities in protein therapeutics. To overcome this challenge, researchers have often needed to perform extensive method development experiments to find the appropriate additives, salt concentrations, and/or organic solvent strengths for accurate quantitation of aggregates and fragments. In this presentation, we will demonstrate how a novel packing material with hydroxy-terminated polyethylene oxide (PEO) bonding and column hardware manufactured to have a hydrophilically modified organosilica surface has led to improved column surface inertness. This technology when paired with an easily prepared buffered mobile phase at physiological pH, is shown to significantly minimize method development. The improved column inertness is demonstrated for separations of protein therapeutics, including monoclonal antibodies, antibody drug conjugates, fusion proteins, and CRISPR Cas proteins along with their aggregates and ribonucleoprotein complexes.
Learning Objectives:
Participants will be able to quickly develop size-exclusion chromatographic methods for biotherapeutics and protein characterization.
Participants will be able to obtain reliable and reproducible results for aggregates, monomer, and fragments species of protein molecules by size-exclusion chromatography.
Participants will be able to use a single buffer for multiple protein and biotherapeutic characterization.
Participants will be able to explore method capabilities for native SEC-MS for CRISPR Cas9 proteins and complexes.
