Therapeutic Antibody Glycan Characterization

Glycosylation significantly affects the effector functions of the immune system, including complement-dependent cytotoxicity (CDC), antibody-dependent cell-mediated cytotoxicity (ADCC) and antibody-dependent cellular phagocytosis (ADCP). This is especially important when one or more of these effector functions are the main mechanism of action. In addition, the carbohydrate structure attached to the monoclonal antibody crystallizable fragment (Fc) affects efficacy and safety, so they are usually classified as key quality attributes. Therefore, the analysis and characterization of glycosylation is an important part of protein structure analysis in the entire development process.

The complex branching and heterogeneous nature of glycans pose significant analytical challenges to their identification and characterization. The coupling of liquid chromatography (LC) and mass spectrometry (MS) has emerged as one of the most powerful tools for the characterization of glycan structures. At Creative Biolabs, we present a streamlined end-to-end analytical platform used for the characterization of glycosylation profile for a therapeutic mAb (IgG1). In this platform, we use new column technology and high-resolution mass spectrometer to simplify the sample preparation program where N-linked glycans were prepared in less than one hour. The analytes were then subjected to HILIC-UPLC/FLR/MS investigations and the resulting data were analyzed to obtain glycosylation profile information of the Fc region for further characterization of IgG1 heterogeneity.

Glycosylation Analysis Services

Broadly speaking, our therapeutic protein glycosylation analysis service can be performed on three levels:

• Intact glycoprotein analysis - MS and other spectroscopic techniques.
• Glycopeptide analysis - HPLC or MS.
• Analysis of released glycans - HPLC/UPLC, and MS.

Fig.1 The top, middle, and bottom levels of glycosylation analysis.^{1, 2}

Sample Data

Fig.2 Glycan profiles of human serum IgG and anti-HER2 monoclonal antibody.^{1, 2}

Fig.3 Determining monosaccharide compositions and linkages using exoglycosidase array digestion of N-glycans.^{1, 2}

For more detailed information about our glycan characterization services, please explore:

• Therapeutic Antibody N-Glycan Analysis Services
• Therapeutic Antibody Glycosylation Site Occupancy Analysis Services
• Therapeutic Antibody Sialic Acid Analysis Services
• Therapeutic Antibody Glycopeptides Analysis Services
• Therapeutic Antibody Intact Released Glycan Analysis Services
• Lectin Microarray-based Glycan Profiling Services
• Glycan Microarray-based Binding Profiles Analysis Services

With extensive experience in glycan characterization, our team can work on custom solutions to meet the unique requirements of each product. With flexible global capabilities, you can provide services where you need it most.

Contact our expert scientists today to discuss how our range of glycan analytical services can improve your unique process.

References

1. Zhang, Peiqing, et al. "Challenges of glycosylation analysis and control: an integrated approach to producing optimal and consistent therapeutic drugs." Drug Discovery Today 21.5 (2016): 740-765.
2. under Open Access License CC BY 4.0, without modification.