Glycosylation Site Profiling Service
Glycosylation site profiling mainly contributes to the illustration of glycan microheterogeneity, which is important for the understanding of glycoprotein biosynthesis and function. As a forward-looking company as well as a market leader in the field of therapeutic antibody development and analysis, Creative Biolabs has successfully accomplished numerous challenging projects in antibody glycan analysis and glycosylation site profiling. Now, we can provide an integrated platform for one-stop glycosylation analysis of your therapeutic antibody candidates. As your best partner, Creative Biolabs is very glad to revolutionize your particular project and promote your success based on the values of quality, timing, and price.
Glycosylation is Necessary for Antibody Property
Therapeutic monoclonal antibodies (mAbs) exhibit heterogeneity in structure and function because of the complex posttranslational modifications, including glycosylation. It has been documented that glycosylation, especially Fc glycosylation, plays a critical role for the determination of antibody physicochemical properties (solubility, viscosity, protein folding, antigen binding, secretion, immunogenicity, etc.) and biological properties [antibody-dependent-cell mediated cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC)]. Therefore, antibody glycan analysis should be determined to produce and develop human mAb drugs with proper function, efficacy, safety, stability, and consistency.
Fig.1 Schematic workflow exemplifying the glycan analysis of a complex glycoprotein.1
Glycosylation Site Profiling Constitute Glycosylation Analysis
The profile of glycosylation sites is an important component of glycosylation analysis, which can supplement already obtained data on protein structure. The site information provides a basis for future detailed characterization regarding the in vivo relevance of the respective site. Furthermore, glycosylation sites may indicate the orientation of transmembrane proteins because glycosylations are commonly encountered at the extracellular domains of proteins. Therefore, glycosylation site analysis is routinely performed on biopharmaceutical products (such as mAbs and glycoproteins) throughout their development and production processes.
Fig.2 Workflow of the glycosylation sites mapping. (Ying, 2017)
Technologies for Glycosylation Site Profiling
Accurate characterization of glycosylation sites is needed for understanding the functions of glycans in mAbs. To determine the site of glycosylation and the glycans on the glycosylation site, first of all, glycoproteins must be cleaved into small glycopeptides. The enzymes trypsin (a highly specific protease) and (non-specific) are widely used in this process. Then, the glycans and peptide sequence on the glycosylation site can be determined by mass spectrometry (MS), which has been the most powerful tool for glycosylation site identification, occupancy measurement, and site-specific glycan structure characterization. At Creative Biolabs, we can provide a series of methods to perform glycosylation site profiling, including but not limited to the followings.
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RP-HPLC-MS
Reversed-phase high-performance liquid chromatography coupled with on-line mass spectrometry (RP-HPLC-MS) can analyze the fractionated peptide and glycopeptides fragments after digestion of a glycoprotein with specific proteases after reduction and alkylation. This method is semi-quantitative because different glycopeptides will ionize differently in a mass spectrometer. -
LC-ESI-MS
Liquid chromatography electrospray ionization mass spectrometry (LC-ESI-MS) is a hyphenated mass spectrometry technique that combines the resolving power of HPLC separation with high mass accuracy of a mass spectrometer. It provides unparalleled sensitivity and selectivity for detection and quantification of site-specific glycan structure. -
HILIC-SPE-MS
Solid-phase extraction using hydrophilic interaction liquid chromatography-mass spectrometry (HILIC-SPE-MS) has become increasingly popular in the glycosylation site mapping because HILIC-SPE-MS is more selective for glycosylated species and is not biased toward particular glycan types. Furthermore, using that technique requires no chemical alteration of glycans. -
ESI/ETD-CID-MS/MS
Electrospray-ionization/Electron transfer dissociation, collision-induced dissociation, tandem mass spectrometry (ESI/ETD-CID-MS/MS) allows for fragmentation of glycan species to be observed predominantly at the glycosidic linkages while the peptide backbone remains intact. CID provides information about the glycan composition.
Workflow
Your protein samples undergo meticulous denaturation, reduction, and alkylation, followed by enzymatic digestion (e.g., trypsin) to generate glycopeptides. This step is critical for isolating the glycosylated fragments.
Specialized techniques, such as hydrophilic interaction liquid chromatography (HILIC) or lectin affinity chromatography, are employed to selectively enrich glycopeptides, enhancing detection sensitivity.
The glycopeptides are then analyzed using high-resolution liquid chromatography-tandem mass spectrometry. This advanced analytical technique precisely identifies glycopeptides and fragments them to determine glycan structures and peptide backbones.
Raw MS data is processed using sophisticated bioinformatics software. Our experts leverage this data to accurately pinpoint glycosylation sites and characterize the specific glycan structures present.
Rigorous quality control checks are performed throughout the process. A comprehensive report is then generated, detailing identified glycosylation sites, associated glycan types, and quantitative data.
Featured Highlights
- Unrivaled Precision: Identifies glycosylation sites and structures with high accuracy.
- Comprehensive Data: In-depth characterization crucial for biologics and vaccine development.
- Accelerated Research: Speed up your project timelines with efficient analysis.
- Expert Interpretation: Use our scientists' extensive knowledge to gain significant insights.
- Enhanced Quality Control: Essential for ensuring consistency and safety of biotherapeutics.
Customer Reviews
"Enhanced batch consistency Using Creative Biolabs' Glycosylation Site Profiling in our study has dramatically enhanced the batch-to-batch consistency of our recombinant protein, giving us confidence for downstream applications while still satisfying crucial regulatory criteria." - June 2024, Mar**er R.
"Precise Vaccine Design The detailed glycosylation site identification provided by Creative Biolabs was instrumental in guiding the design of our novel vaccine antigen, confirming critical glycosylation for optimal immunogenicity." - April 2025, Jo**n S.
"Comprehensive Biosimilar Analysis Creative Biolabs' service offered a truly comprehensive comparative analysis of glycosylation profiles, which was essential for demonstrating similarity for our biosimilar development program. The data quality was exceptional." - February 2025, Sam**h W.
FAQs
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Q1: What types of samples are suitable for Creative Biolabs' Glycosylation Site Profiling Service?
A1: Our service is highly versatile and can accommodate a wide range of samples, including purified proteins, antibodies, recombinant proteins, cell lysates, and even complex biological fluids. We encourage you to discuss your specific sample type with our specialists for tailored advice.
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Q2: What is the typical turnaround time for a Glycosylation Site Profiling project?
A2: While the exact timeline can vary depending on sample complexity and project scope, most projects are completed within 4 to 8 weeks. We prioritize efficient processing without compromising data quality. For a more precise estimate, please reach out to our team.
Related Services
To further assist your research and development needs, Creative Biolabs offers a spectrum of ancillary services:
- Glycan Sequencing: For detailed structural elucidation of individual glycans.
- Glycoform Analysis: To quantify the distribution of different glycoforms of your protein.
- Monosaccharide Analysis: To determine the monosaccharide composition of your glycans.
Why Choose Us
Creative Biolabs is your top choice for negotiating the complexities of biopharmaceutical development. Our advanced Glycosylation Site Profiling Service, backed by unparalleled expertise and cutting-edge technology, provides the precise insights you need to accelerate discovery, ensure product safety, and enhance therapeutic efficacy. We're committed to delivering high-quality, actionable data that drives your success. Please feel free to reach out to us. We're eager to discuss your specific research requirements and demonstrate how our expertise can directly benefit your goals.
References
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Mastrangeli, Renato, Abhijeet Satwekar, and Horst Bierau. "Innovative Metrics for Reporting and Comparing the Glycan Structural Profile in Biotherapeutics." Molecules 28.8 (2023): 3304.
Distributed under Open Access License CC BY 4.0, without modification. -
Ying, Jiezheng, et al. "Mapping the N-linked glycosites of rice (Oryza sativa L.) germinating embryos." PLoS One 12.3 (2017): e0173853.
Distributed under Open Access License CC BY 4.0, without modification.
For Research Use Only.