Development of a Multi-Product SE-UHPLC Method for the Determination of Size-Variants in Bispecific Antibody Formats

Elevated levels of aggregates in protein therapeutics are associated with an increased risk of triggering adverse immune responses in patients. It is thus of paramount importance to closely monitor these product-related impurities by the most suitable analytical tools. Size exclusion-high performance liquid chromatography (SE-HPLC) is the preferred method applied in quality control laboratories for the quantitative analysis of aggregates (high-molecular-weight forms), the antibody monomer itself, as well as low-molecular-weight forms. Size-exclusion ultrahigh-performance liquid chromatography (SE-UHPLC) using new columns with packings of particles [less than or equal to]3 [micro]m provides significant gains in chromatographic efficiency by a higher resolution, enhanced selectivity, and shorter run times. Here, we present a multi-product SE-UHPLC platform method for the characterization of bispecific monoclonal antibody formats with superior separation characteristics compared to our existing SE-HPLC method, which can be readily applied for process optimization, stability testing, and release analytics.
Biotherapeutics and, more specifically, monoclonal antibodies (mAbs) rank among the fastest growing and most promising classes of drugs in basic research, as well as in clinical settings for various life-threatening [...]