Workflow For High-Throughput Glycoprofiling Of Rituximab

Careful and thorough characterization of therapeutic mAbs is essential for ensuring drug safety and efficacy. mAbs are typically manufactured in mammalian host cell lines in bioreactors, generating a large number of heterogeneous drug molecules. Establishing a number of critical quality attributes (CQAs) for each mAb and demonstrating that production batches are within acceptable limits are requirements for both innovator and biosimilar therapeutics.

In many cases, the characterization of an antibody-based drug is performed using a specific chromatographic technique (e.g., size exclusion, reversed phase or hydrophilic interaction liquid chromatography, respectively – SEC, RP or HILIC) coupled with mass spectrometry (MS). This combination allows for different types of analyses to be carried out, e.g., accurate mass measurement of the intact mAb and subunits, peptide mapping, and the determination of post-translation modifications such as glycosylation, oxidation, and deamidation.

Discover a complete SEC-MS workflow that has been developed to enable rapid glycoprofiling of monoclonal antibodies (mAbs) in cell culture supernatants. In detail, this application note includes:

• Antibody (mAb) purification using immobilized protein A
• Antibody reduction procedure
• Mass spectrometer calibration
• System suitability test utilizing a recombinant human monoclonal antibody reference
• Size exclusion chromatography - mass spectrometry (SEC-MS) method for sample separation and analysis