Glycosylation of proteins is one of the most common post translation modifications (PTMs) and also a critical quality attribute of biopharmaceutical products. Glyco-proteins contain complex sugar moieties whose presence, absence, sites of attachment, and relative abundance can have a major impact on the efficacy, stability and activity of proteins and monoclonal antibodies (mAbs). Hence, it is essential to determine the glycan structure and heterogeneity, through glycosylation analysis.
A full glycosylation analysis includes the number, position and type of glycosylation in your protein or mAb. The analysis is especially useful for comparing different batches, originator versus biosimilar and process development samples.
Select from the following service:
Glycosylation analysis can be done at different levels. It can also be combined to achieve a full glycan characterization.
Service 1: Separation and exact molecular weight of protein/mAb glycoforms
- Firstly, determination of exact molecular weight of glycoprotein by LC-MS (HILIC separation and online LC-MS).
- Then, determination of exact molecular weight of deglycosylated protein by LC-MS (HILIC separation and online LC-MS).
Service 2: Separation and exact molecular weight of mAb sub-units
- IdeS digestion and reduction to sub-units.
- HILIC separation of both LC, Fd’ and single chain Fc (scFc) glycoforms.
- Online LC-MS to determine the mass of each sub-unit.
Service 3: Glyco-peptide mapping
- Glycoprotein digest with one of more proteases (trypsin or Lys-C) and analyzed by LC-MS/MS.
- Optional: Collection of glycopeptide fractions and offline MALDI TOF/TOF analysis.
- Identification of glycopeptides by searching against drug substance sequence and glycostructure database.
Service 4: Released N-glycan analysis
- PNGase F treatment to release N-linked glycans.
- Fluorescent labelling using Waters RapiFluor Glycan labelling kit.
- Separation of glycans by HILIC chromatography, detection by fluorescence HPLC and mass spectrometry.
- Optional: Collection of the glycan fractions for offline MALDI TOF/TOF analysis.
- Identification of both glycan profile and type, by FLD-trace and glycan mass.