Services Intact mass analysis and peptide mapping

Intact mass analysis and peptide mapping services

This page has been reviewed and verified by Thomas Kofoed, PhD in Organic Chemistry

By utilizing mass spectrometry (LC-MS), you get a fast, reproducible, quantitative evaluation of product-specific attributes and process-related impurities. Our CRO services include intact mass analysis and peptide mapping to provide a detailed characterization of your drug substance (DS) for use in process and control strategy optimization. Furthermore, you’ll get a thorough overview of your DS’s stability, function, and purity.

Reproducible, quantitative intact mass and peptide mapping enable:

Detailed product and process understanding

Optimize the production process and evaluate the impact of formulation and manufacturing changes on product quality

Ensure the quality of your product throughout its lifecycle

Develop control strategies for monitoring product characteristics affecting product quality, and reduce the risk of variability in the final product

Faster regulatory approval

Increase transparency and understanding of your product’s chemistry, manufacturing, and controls (CMC) for expedited processing by regulatory authorities

Protein structure characterization and quantification of post-translational modifications

The ICH Q6B guidelines call for thorough protein structure characterization of biologics and assessment of drug substance heterogeneity. You must demonstrate protein and batch consistency during drug development, following process or manufacturing changes, and after up-scaling or contract manufacturing organization (CMO) tech transfer. Early, pre-clinical CQA analysis helps prevent delays due to stability issues or other challenges during the clinical trial stages.

Intact mass analysis and peptide mapping services using LC-MS are highly sensitive and selective techniques for determining structural characterization and post-translational modifications (PTMs), which may affect the quality and efficacy of your final drug product. LC-MS data provides strong evidence of correct expression of specific proteins, protein impurity clearance, and product stability for regulatory submissions and product release testing.

Glycosylation is a typical PTM whose presence or absence, sites of attachment, and relative abundance can significantly impact the efficacy, stability, and activity of proteins and monoclonal antibodies (mAbs). We can help you compare batches, originator and biosimilar, or your process development samples using intact mass analysis and/or a complete glycosylation characterization - including N-glycan analysis.

Our LC-MS-based quantitative peptide mapping services also allow a thorough characterization of other PTMs, such as deamidation, oxidation, methylation, and pyro-Glu.

Do you know your product’s disulfide bonds and free cysteines?

For many proteins and mAbs, correct disulfide linkage is critical for optimal biologic function. Product stability and activity issues often correlate with an abnormal pattern of disulfide bonds which affect the protein conformation and structure.

For this reason, the ICH Q6B guidelines call for analyzing disulfide bonds and free cysteines in biopharmaceuticals.

Using LC-MS, we perform protein mapping to determine the number and expected positions of disulfide bridges and free sulfhydryl groups in both non-reduced and reduced proteins and mAbs.

Does your documentation of CQAs fulfill FDA requirements?

To avoid significant setbacks during regulatory approval, you must have a complete picture of your protein product’s CQAs and use the optimal analytical assays for each attribute to document compliance with regulatory requirements.

Combining different types of chromatography with our mass spectrometry services for intact mass analysis and quantitative peptide mapping provides the CMC department with the most comprehensive data available for documentation.

Overview of our analyses according to ICH Q8B

Quality attribute (QA)		Analytical Service
Primary sequence	Molecular mass Amino acid sequence coverage Free cysteines	LC-MS intact mass Peptide mapping (1-3 different enzymatic digests) MS-based method
Glycosylation profile	From intact mAb Map of N-glycosylation	RPLC MS Peptide mapping Release N-glycan analysis
Product heterogeneity, related substances, and impurities	N- and C-terminal modifications Charge and size heterogeneity	IEX MS or SEC/RPLC MS (incl. de-glycosylation) Peptide mapping IEX MS
Chemical modifications	Position of glycation	Peptide mapping (trypsin digest plus extra if needed) IEX MS with de-glycosylation Alternatively, RPLC MS or SEC MS with de-glycosylation
Secondary structures	Disulfide linkages	Peptide mapping (non-reduced)

Client stories

Video: GMP-validated HCP analysis based on LC-MS

The world’s first mass-spectrometry-based analysis of HCP impurities under GMP conditions

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Video: Analyzing unexplainable HPLC peaks or shoulders

Using an IEX-MS setup as an essential part of quality control and documentation, especially when experiencing new peaks or shoulders

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Stability study – an important part of biologics license application

Our client needed a stability study of protein degradation products for regulatory documentation

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Characterization program for therapeutic mAbs

“We identified a scale-up problem and now use the analysis to test the quality of all our batches”

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Antibody characterization for cancer drug

High-throughput antibody characterization service

GTP Bioways develops a massive number of high-quality mAbs with Alphalyse’s LC-MS analysis for characterization

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Optimized HPLC analysis of peptides for clinical trials

DTA Consulting wanted to improve their client's HPLC setup for GMP production of a peptide with high stability.

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Typical project process

You typically work with
these MS and protein experts:

Marcella Nunes de Melo-Braga

PhD in Protein Chemistry and Proteomics

Stine Thyssen

Head of Analytical Characterization Dpt

Maša Babović

PhD in Biochemistry and Molecular Biology

Project scope
We like to start with an online meeting to learn more about your project. Based on your needs and details about your samples, you will receive a draft proposal outlining the suggested analyses and expected timeframe.
Samples
After signing the final project proposal, we will contact you for information about shipping samples. We will inform you of an estimated report delivery date as soon as we receive your samples.
Execution
A project leader will oversee the project and send you status updates by email at regular intervals.
The analysis varies according to the project but typically includes:
- Intact mass analysis performed by LC-UV-ESI MS.
- Reduction/alkylation, protease digestion, and LC-ESI MS/MS peptide.
- Disulfide bond mapping by non-reduced peptide mapping analysis.
- Quantification of major post-translational modifications.
- N-glycan analysis.
Results
You will receive the analysis report by email. Depending on the project, it includes:
- Objectives, description of analytical procedure, results, and conclusions.
- Selected raw data, e.g., excel sheets. Additional raw data may be provided to you upon request.
Follow up
Upon project completion, your team is invited to review the results at an online meeting.

What clients say

"The collaboration with Alphalyse was superb! They quickly presented an optimized method that provided excellent data and separated product-related impurities. The contribution made it possible to keep our milestones" Dorrit Andersen, Regulatory Affairs Consultant DTA Consulting, Denmark

"They handled the project professionally and rapidly, and the report was very well written, clearly explaining the findings."

Kristiina Hyvärinen, Director QC, viral products
Targovax ASA, Finland

"We enjoy collaborating with Alphalyse as part of our optimization of manufacturing processes. Not only do we gain access to their hands, but we also get to pick their brains for mass spectrometry knowledge."

Torben Lund-Hansen, PhD, SVP
Head of Technical Operations
Y-mAbs Therapeutics Inc., USA

"We are very pleased with the work of Alphalyse because they provide us with a high-quality antibody characterization service. Most importantly, we can ship them hundreds of samples at once and always receive the analytic results shortly after. "

Head of CMC, C&GT Division
GTP Bioways, France

"We received a very professional report. It included an excellent overview of the peaks and their identity in the different batches and degradation samples. With the documentation from the study, we finally had everything we needed to send in the Biologic drugs license application (BLA) for FDA approval."

CMC Project Leader, R&D
UK biopharma company

"With the results from the extensive characterization, we identified a scale-up problem and could optimize the process to increase mAb stability. Therefore, we now use the analysis on an ongoing basis to test the quality of all our batches"

Director, Protein and Analytical Chemistry
Research-based pharmaceutical company

“I found the group at Alphalyse knowledgeable, easy to work with, and helpful in
planning the study”

VP CMC and Quality
US Biotech

“Extremely professional service, high-level quality of the results, and excellent communication”

Program Manager
European Biotech

Knowledge center

FAQ

Why combine intact mass analysis and peptide mapping services for finding critical quality attributes?

Intact mass analysis is superior for identifying and quantifying different modifications in your intact drug molecule, whereas peptide mapping locates each modification in detail. Combining the two analyses allows e.g., detection that a molecule has 10 % oxidation using intact mass analysis, followed by peptide mapping to identify the exact site of the oxidations.

Why should I identify the Critical Quality Attributes (CQAs) in my drug molecule?

Knowing the CQAs of your biopharmaceutical can help you anticipate challenges and issues that may influence the stability and activity of your drug product. Monitoring CQAs is essential to ensure consistency of the manufacturing process and comparability of DS batches, as well as for regulatory documentation.

What are the typical CQAs of protein biologics?

Typical CQAs can be specific glycoforms, charge variants (e.g., deamidation), easily oxidized sites, and aggregations.

Resources

Instrumentation

Talk to us

Whatever protein-related challenge or question you may have, we would love to help. Our experts can help you decide on the best analytical approach for your project by email or online meeting - providing advice without obligation.

Intact mass analysis and peptide mapping services

Reproducible, quantitative intact mass and peptide mapping enable:

Detailed product and process understanding

Ensure the quality of your product throughout its lifecycle

Faster regulatory approval

Protein structure characterization and quantification of post-translational modifications

Do you know your product’s disulfide bonds and free cysteines?

Does your documentation of CQAs fulfill FDA requirements?

Overview of our analyses according to ICH Q8B

Client stories

Video: GMP-validated HCP analysis based on LC-MS

Video: Analyzing unexplainable HPLC peaks or shoulders

Stability study – an important part of biologics license application

Characterization program for therapeutic mAbs

High-throughput antibody characterization service

Optimized HPLC analysis of peptides for clinical trials

Typical project process

Project scope

Samples

Execution

Results

Follow up

What clients say

Knowledge center

FAQ

Why combine intact mass analysis and peptide mapping services for finding critical quality attributes?

Why should I identify the Critical Quality Attributes (CQAs) in my drug molecule?

What are the typical CQAs of protein biologics?

Resources

Videos:

Client cases:

Talk to us