Wide range of biological samples
Protein glycosylation is a non-template driven postranslational modification that not only leads to considerable product heterogeneity in recombinant protein expression but also effects efficacy, circulatory half-life and immunogenicity of the biopharmaceutical drugs. For this reason protein glycosylation has become a critical quality attribute in therapeutic protein production and requires a continuous analysis through the entire development and production cycle.
We offer custom glycan analysis services for a wide range of biological samples
- Serum antibodies and/or full serum glycan profile
- Saliva, urinary glycoproteins
- Recombinant purified proteins
- Protein extracts
- Exosomes
- Plant proteins
- Cell surface glycans
Addressing your needs
We offer you a stepwise glycan analysis service of increasing complexity.
Glycan analysis can be very complex and we will help you choose the correct strategy to obtain the information your really need. We offer a constantly updated portfolio of glycan analysis modules of increasing complexity including glycan profiling, structure assignment and glycosylation site ocupancy for O- and N-glycosylation. Our analytical services have been used in biomarker discovery and validation studies, comparison of glycan profiles for different production cell lines, in allergy research, process optimization and early stage biopharmaceutical development.
Routine Glycan Analysis Services
Carboquant N-glycan analysis
Absolute and relative quantification of N-glycans by MALDI-TOF MS with 13C labeled internal glycan standards
- Addition of 8-10 13C labeled internal standards
- Protein denaturation
- Enzymatic removal of N-glycans
- Glycan analysis by MALDI-TOF MS
- Tentative peak assignment and quantification with dedicated quantification software
- Report
Profiling of permethylated N-glycans by MALDI-TOF MS
Semiquantitative analysis of N-glycan structures by MALDI-TOF MS after derivatisation with methyliodide to increase signal intensity stabilize labile sialic acid residues.
- Protein denaturation
- Enzymatic removal of N-glycans
- Labeling (permethylation)
- Glycan analysis by MALDI-TOF MS
- Tentative peak assignment and quantification
- Report
N-glycan profiling by UPLC-FLD/MS
Profiling of procainamide labeled N-glycans by fluorescence and mass detection. Relative quantification of glycan abundance by integration of corresponding fluorescence peak area. Individual quantification of chromatographically resolved isomeric structures possible.
- Protein denaturation
- Enzymatic removal of N-glycans (PNGAse F)
- Fluorescent labeling of N-glycans with procainamide
- Glycan analysis by UPLC-FLD/MS
- Tentative peak assignment and relative quantification of glycans
- Report
Sialylated N-glycan profiling by MALDI-TOF MS
Focus on the differentiation of α-2,3 and α-2,6 sialic acid linkages and NeuNAc and NeuNGc residues in the N-glycan profile.
- Protein denaturation
- Enzymatic removal of N-glycans (PNGAse F)
- Enrichment of sialylated glycans
- Stabilization of sialic acid residues by esterification/lactonisation
- N-glycan analysis by MALDI-TOF MS
- Peak assignment and relative quantification of structures
- Report
O-glycan profiling
Analysis of permethylated O-glycans by MALDI-TOF MS
- Protein denaturation
- Enzymatic removal of N-glycans (PNGAse F)
- Removal of O-glycans by β-elimination with NaOH
- Sample clean-up
- Permethylation of isolated O-glycans
- Analysis by MALDI-TOF MS
- Peak assignment and relative quantification of structures
- Report
Relative sialic acid quantification (NeuNAc/NeuGc ratio)
Quantitative analysis of sialic acids following Waters Corp. technical note “DMB-labeled Sialic acid analysis Using UPLC based BEH C18 columns”
- Mild acetic hydrolysis of sialic acid residues
- Fluorescent labeling with DMB
- Chromatographic separation of sialic acid derivatives by reverse phase UPLC and detection via fluorescence
- Peak assignment by comparison with external standards and mass spectrometry
- Report
2nd level glycan assignment
The correct asignment of isomeric structures can require a more sophiscated analytical approach that includes
- Analysis of diagnostic fragments generated in LC-MS/MS or MALDI-TOF MS workflows
- Exoglycosidase based glycan sequencing coupled with LC-MS to confirm monosaccharide type and the stereochemistry of glycosidic linkages (e.g. alpha or beta-linked galactose residues)
- Use of in house reference standards (SIL-standards and fluorescently labeled structures)
We offer UPLC-FLD/MS of 2-AB, 2-AA or procainamide labeled glycans profiling service of cell, tissue or glycoprotein samples, and MALDI-TOF MS based glycan structure identification, including α-2,3/ α-2,6 differential sialylation.
Custom glycan analysis projects
Please inquire for any other glycan analysis project not included in our routine services. This includes
- Plant N- and O-glycosylation
- Polysaccharides
- Glycolipids
- Milk oligosaccharides
- N- and O-glycosylation site analysis
- Development of customized sample preparation and glycan enrichment protocols
- lectin-base glycan analysis (see also lectin microarray services)
- general glycan quantification
- Preparative isolation of glycans for analytical purposes
- use of other fluorescent labels like RapiFluor, 2AA, 2AB, etc.
Mabsolute- Absolute quantification of IgG glycans
Our CarboQuant HTS glycan quantification service is based on isotopic dilution and applied to the absolute quantification of N-glycans in IgG type monoclonal antibodies by MALDI-TOF MS, both in low and high-throughput (up to 96 samples).
- Glycoanalysis of biologics
- Absolute quantification of biosimilar glycosylation
- For analysis of other glycoproteins please inquire
Why our service?
- Only service that currently provides absolute Glycan Quantification (pmol).
- Glycan quantification with internal standards (not based on quantification of attached fluorophores).
- Simple sample preparation and data acquisition.
Report includes
- Absolute Glycan Quantification (pmol).
- Relative Glycan Quantification (%).
- Absolute Monosaccharide Quantification (pmol).
- Relative Monosaccharide Quantification (%).
- Total Glycan amount (pmol).
Sample requirements
- Minimum required sample amount is 50 μg of human IgG type antibodies expressed in CHO cell lines.
- optimal sample amount 50-500 μg of IgG type antibody expressed in CHO cell lines.
- For optimal sample condition ship at 4 ºC or – 20 ºC.