Biotherapeutics such as monoclonal antibodies (mAbs) are frequently glycosylated, and the structure of attached N-glycans can affect immunogenicity, pharmacokinetics and pharmacodynamics. This can make glycosylation a critical quality attribute (CQA), making characterization of N-glycans an essential part of the development process. We present complete N-glycan quantitation workflow solutions using liquid chromatography (LC) with fluorescence detection (FLD) or mass spectrometry (MS) for four levels of biotherapeutic analysis: intact mass, mAb subunits, glycopeptides, and released glycans. Released N-glycans are labeled with InstantPC (a glycan dye that provides high FLD and MS signal) or 2-AB (a traditionally-used label) prior to separation by hydrophilic interaction liquid chromatography (UHPLC-HILIC). Sialylation of biotherapeutics can also be important, and we present a plate-based streamlined workflow for quantitation of total sialic acid.
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