Monosaccharide Analysis Service
Monosaccharide composition analysis is routinely performed on biopharmaceutical products [such as monoclonal antibodies (mAbs) and glycoproteins] throughout their development and production processes to ensure proper function, efficacy, and safety, stability, serum half-life, as well as for quality control and release purposes. As the long-term pioneer as well as market leader in the field of therapeutic antibody development, Creative Biolabs has successfully developed a versatile antibody glycan analysis platform to illustrate antibody monosaccharide composition and screen the changes in glycosylation. 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 Alters the Physicochemical Property of mAbs
Glycosylation represents a major post-translational modification of monoclonal antibodies (mAbs) and glycoproteins that are involved in various biological processes, including transcription, differentiation, apoptosis, cell adhesion, receptor-ligand binding, as well as oncogenic transformation and immune response. Particularly, these glycosylated mAbs and proteins are one of the fastest growing therapeutic drugs and their activity, stability, serum half-life and adverse reactions are largely determined by glycosylation. Glycosylation is, therefore, one of the critical quality attributes that must be rigorously characterized and monitored.
Fig.1 Summary of IgG N-glycan present on the variable region (Fab) and the constant heavy chain tail region (Fc) with associated immunologic functions.1
Importance of Monosaccharide Analysis in mAb Characterization
Monosaccharide composition analysis can screen for changes in glycosylation as well as quantify the amounts of individual monosaccharides, most often fucose, galactosamine, glucosamine, galactose, glucose, and mannose. Most mammalian glycoproteins contain three major types of glycans: N-linked, O-linked and glycosylphosphatidylinositol (GPI) lipid anchors, which consist of one or more monosaccharide units. For therapeutic mAbs, monosaccharide composition analysis is extremely important due to its important role in affecting both antibody characteristics (e.g., antigenicity, stability, folding, serum half-life, and antigen binding affinity) and effector functions [e.g., antibody-dependent-cell mediated cytotoxicity (ADCC), complement-dependent cytotoxicity (CDC)]. Especially, beyond profiling changes in total glycan composition, monosaccharide analysis can also be used as an exploratory technology to identify various monosaccharide modifications, including phosphorylation and sulfation.
Main Methods of Monosaccharide Analysis
To support researchers in glycoprotein engineering, we provide a series of technologies for monosaccharide analysis, including but not limited to HPAEC-PAD, HPLC, and MS.
- HPAEC-PAD
High-performance anion exchange chromatography coupled with pulsed amperometric detection (HPAEC-PAD) is a classic procedure used for the analysis of monosaccharides released from a glycoprotein. With high selectivity and specificity, HPAEC-PAD permits direct analysis of nonderivatized monosaccharides, providing not only the total glycosylation status of a protein, but also the amounts of specific monosaccharides. Particularly, HPAEC-PAD analyzes the carbohydrate species without the need for derivatization. Generally, samples are acid hydrolyzed to release the monosaccharides, lyophilized, dissolved in water, and then analyzed by HPAEC-PAD. This specific method can not only provide details about fucosylation, an indication of the presence of O-linked glycans, but also determine mannose content as an indication of high-mannose N-linked glycans.
- HPLC-Based Monosaccharide Analysis
High-performance liquid chromatography (usually with fluorescence detector) is another common method for the determination of two most abundant sialic acids (Neu5Ac and Neu5Gc) by derivatization. During this analysis process, 1,2-diamino-4,5-methylenedioxybenzene dihydrochloride (DMB) is typically used to modify the sialic acids after release from glycoprotein to provide a fluorescent product. HPLC allows separation and detection of a wider variety of modified sialic acids. Highly efficient hydrophilic interaction chromatography (HILIC)-based ultra-performance liquid chromatography (UPLC) is emerging as a powerful tool for monosaccharide composition analysis.
- Mass Spectrometry (MS)
Mass spectrometry is a versatile technique that has increasingly been used in monosaccharide analysis due to its sensitivity, potential for high throughput analysis, and the capability for tolerating diverse biological mixtures. Separation techniques such as liquid chromatography (LC) and ion mobility are typically used in conjunction with mass spectrometry to analyze the antibody monosaccharide with more accuracy.
Required Starting Materials
- Purified Glycoproteins or Therapeutic Antibodies: These are ideal for direct analysis of their specific glycan components.
- Cell Lysates or Culture Supernatants: Useful for studying cellular glycosylation patterns or secreted glycoconjugates, providing insights into cellular processes.
- Glycosylated Drug Candidates or Vaccine Components: Essential for characterization during early-stage development and for ensuring quality assurance throughout the manufacturing process.
Our Monosaccharide Analysis Workflow
Step1. Sample Preparation
This crucial initial step involves meticulous purification, concentration, and often desalting of your sample to remove any interfering substances. The outcome is a highly purified and concentrated sample, optimized for subsequent analytical stages, ensuring the highest data quality.
Step2. Glycan Release (if applicable)
For complex glycoconjugates where glycans are covalently attached, these carbohydrate chains are enzymatically or chemically released from the protein backbone. This yields free glycans, simplifying downstream analysis and allowing for a focused examination of the carbohydrate moiety.
Step3. Hydrolysis
The released glycans or intact glycoconjugates are then subjected to controlled acid hydrolysis. This process breaks them down into their individual constituent monosaccharide units. This step is vital as it provides the isolated building blocks for specific identification and quantification.
Step4. Derivatization
Each individual monosaccharide unit obtained from hydrolysis undergoes a precise chemical modification, known as derivatization. This process significantly enhances their detection sensitivity and improves their separation efficiency during chromatographic analysis. The result is a set of stable, highly detectable derivatives for each monosaccharide.
Step5. Chromatographic Separation & Detection
The derivatized monosaccharides are separated with exceptional resolution using advanced high-performance liquid chromatography (HPLC) or gas chromatography (GC). Following separation, they are detected by sensitive techniques such as fluorescence, ultraviolet (UV) absorption, or mass spectrometry (MS). This stage generates distinct peaks for each monosaccharide, allowing for both qualitative identification and accurate quantitative measurement.
Step6. Data Analysis & Interpretation
The raw chromatographic and spectrometric data are then rigorously processed, quantified, and interpreted by our team of expert glycobiologists. This final step culminates in a comprehensive report detailing the qualitative identification and precise quantitative composition of the monosaccharides in your original sample, complete with biological insights.
Service Highlights
- Unparalleled Precision: Achieve highly accurate and reliable characterization of glycan components.
- Accurate Quantification: Obtain precise quantification of individual monosaccharides, crucial for detailed structural and functional analysis.
- Critical Structural Insights: Gain essential understanding of protein function and stability through detailed carbohydrate profiling.
- Rigorous Quality Control: An indispensable tool for stringent quality control and comparability studies in biopharmaceutical manufacturing.
- Regulatory Compliance: Ensures batch-to-batch consistency and aids in meeting crucial regulatory requirements.
- Comprehensive & Efficient: Our approach provides a thorough analysis with streamlined processes.
- Detailed Carbohydrate Profiles: Obtain comprehensive and actionable insights into the carbohydrate composition of your samples.
Customer Reviews
Precise Composition (April 2025), Mar*** Z
Using Creative Biolabs' Monosaccharide Analysis in our research has significantly improved our ability to confirm the exact carbohydrate composition of our recombinant proteins, which is critical for downstream functional assays and stability testing.
Invaluable QC (January 2025), Vic**** P
The detailed monosaccharide profiles provided by Creative Biolabs have significantly facilitated our quality control processes for vaccine batches. The data allowed us to rapidly detect and correct minor glycosylation shifts, ensuring consistency and safety across production runs.
Enhanced Comparability (June 2025), Sar*** L
Creative Biolabs' Monosaccharide Analysis proved invaluable for our biosimilar comparability studies. The clear, quantitative data on monosaccharide ratios helped us confidently demonstrate structural similarity to our reference product, streamlining our regulatory pathway.
FAQs
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Q1: What types of samples can be analyzed using Creative Biolabs' Monosaccharide Analysis service?
A1: We are equipped to analyze a wide range of samples, including purified glycoproteins, therapeutic antibodies, cell lysates, vaccine components, and other complex glycoconjugates. If you have a unique sample type, please reach out to our team to discuss its suitability and optimal preparation.
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Q2: How does Creative Biolabs ensure the accuracy and reliability of its Monosaccharide Analysis results?
A2: Our commitment to accuracy is unwavering. We utilize state-of-the-art chromatographic and mass spectrometric platforms, employ rigorous quality control measures, and adhere to strict standard operating procedures. All analyses are performed by highly experienced scientists, ensuring precise and reliable data for your project's critical needs.
How to Contact Us
Creative Biolabs is your trusted partner for comprehensive monosaccharide analysis, providing the precision and expertise required to navigate the complexities of glycobiology in drug discovery and development. Our advanced analytical capabilities and commitment to scientific excellence ensure you receive reliable, actionable data for your most challenging projects, accelerating your path to success. Please click the link to get in touch with us for more details.
Reference
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Haslund-Gourley, Benjamin S., Brian Wigdahl, and Mary Ann Comunale. "IgG N-glycan signatures as potential diagnostic and prognostic biomarkers." Diagnostics 13.6 (2023): 1016.
Distributed under Open Access License CC BY 4.0, without modification.
For Research Use Only.