Peptide Mapping

Biopharmaceutical peptide mapping

Peptide mapping is a critical workflow in biotherapeutic protein characterization and is essential for elucidating the primary amino acid structure of proteins. For recombinant protein pharmaceuticals, such as monoclonal antibodies (mAbs) and antibody-drug conjugates (ADCs), peptide mapping is used for proof of identity, primary structural characterization and quality assurance/quality control (QA/QC). Global regulatory agencies, including US Food and Drug Administration (US FDA) and European Medicines Agency (EMA), look to harmonized guidelines from the International Council for Harmonisation (ICH). ICH Q6B  covers the test procedures and acceptance criteria for biologic drug products, and specifies the use of peptide mapping as a critical quality test procedure for drug characterization used to confirm desired product structure for lot release purposes.

Comprehensive peptide maps, simplified

In order to generate a peptide map, the therapeutic protein must first be digested into its constituent peptides via a chemical or enzymatic reaction. Robust separation and identification of the resultant peptides then provides insight into a protein’s full sequence information; displaying each amino acid component and the surrounding amino acid microenvironment, including disulfide linkage information. Structural characterization at this level highlights post translational modifications (PTMs) such as site-specific glycosylation, amino acid substitutions (sequence variants) and/or truncations which may result from erroneous transcription of complementary DNA. Within a bioproduction environment, peptide mapping is necessary for manufacturing process monitoring and QC. It facilitates product comparability testing, which is necessary to identify any product-related impurities, such as deamidation and/or oxidation following any formulation, manufacturing process or storage change.

Due to its complexity and inherent variability, peptide mapping is generally performed in a comparative manner; for example, biosimilars would be compared to a reference or control substance, such as the innovator biologic, in a side-by-side experiment. An in-depth analysis is then required to identify minor and even isobaric differences in protein primary structure. The modern biopharmaceutical and protein research laboratory is tasked with providing high quality analytical results, often in high-throughput, regulated environments. Some technologies currently employed for biopharmaceutical peptide mapping are subject to high levels of irreproducibility, poor sensitivity, and high levels of time-consuming manual work—with protracted methodologies that are not amenable to automation and often require 24 hours to achieve full protein digestion.

This variability impacts data confidence. Moreover, it increases potential for introduction of non-product related artifacts during manual sample handling.

The most recent advances in protein sample preparation chemistries, ultra-high performance liquid chromatography (UHPLC), mass spectrometry (MS) hardware and intuitive software facilitate the generation of comprehensive, confident peptide maps.

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Type Title
AppsLab AppsLab Peptide Mapping eWorkflows
Application Note Peptide Mapping of Challenging Monoclonal Antibodies
Application Brief High Resolution, High-Precision Peptide Mapping of a Monoclonal Antibody Digest
Application Note Comparison of Alternative Approaches to Trypsin Protein Digestion for Reproducible and Efficient Peptide Mapping Analysis of Monoclonal Antibodies
Application Note High-throughput Peptide Mapping of Trastuzumab using a Tandem LC-MS Workflow
Application Note Automated Biopharmaceutical Protein Digestion Optimization for Peptide Mapping Method Development
Application Note High-precision, automated peptide mapping of proteins
Application Note Fast Digestion Method Optimization Using Innovative Trypsin Technology
Application Note High-Throughput Peptide Mapping with the Vanquish UHPLC System and the Q Exactive HF Mass Spectrometer
Application Note Impact on Tryptic Digestion when Detergents and Chaotropes are Essential: A Case Study with Ribonuclease A
Application Note Improvement in Speed and Reproducibility of Protein Digestion and Peptide Quantitation, Utilizing Novel Sample Preparation Technology in a Full Solution Workflow
Application Note Increased Efficiency of Biomolecule Identification by Optimization of Trypsin Digestion Buffers
Application Note Increased Long-term Stability of Peptide Mapping Using the Vanquish UHPLC System
Application Note LC-UV-MS Peptide Mapping Development for Easy Transfer to LC-UV QA/QC
Application Note Increasing Sensitivity in HCP Analysis: from Simplified Sample Preparation to Reliable Results
Application Note Reliable Results in Peptide Mapping Using the Vanquish Flex UHPLC System
Application Note Selecting Buffers to Remove Uncertainty in Tryptic Digestion
Application Note HILIC – an alternative separation technique for glycopeptides
Application Note Investigating Process-Related Post-Translational Modifications in NISTmAb RM 8671 Using High-Throughput Peptide Mapping Analysis
Article Developments in Peptide Mapping Technology for the Biopharmaceutical Industry
Article Column Temperature Control in Peptide Mapping. Maximizing Peak Capacity by Keeping Your Column Hot the Right Way.
Article High-Throughput, High-Resolution Peptide Maps
Blog Admiring the Architecture of the Building Blocks of Life
Blog Ahoy! Do I Need Satellite Navigation for my Peptide Map?
Blog Finding HOPE Using Peptide Mapping?
Blog Not All Who Wander Are Lost
Blog What’s faster? Digesting a protein, or reading this blog?
Blog BioPharMoore: Unravelling Biopharmaceutical Protein Digestion
Brochure Peptide Mapping Workflows: Upgrade Your Maps
Brochure Vanquish Flex UHPLC System: Built for BioPharma
Brochure BioPharma Finder Software - Brochure
Brochure Q Exactive BioPharma
eBook Putting Peptides on the Map
eBook Simpler, Faster and More Reproducible Protein Digestion
Online Tool SMART Digest Kits - Digest Time Calculator
Poster Differentiate Minor Difference of Protein Structure in Biosimilar and Reference Products Using High-Resolution Orbitrap LC-MS/MS
Poster Note Characterization of Monoclonal Antibodies and ADCs using a Benchtop Orbitrap Mass Spectrometer
Poster Note Comprehensive Sequence and Post-translational Modifications Analysis of Monoclonal Antibody by Flash Digest and LC-High Resolution MS
Poster Note Degradation Profiling of a Monoclonal Antibody Using Multiple Fragmentation Techniques and a Novel Peptide Mapping Software
Poster Note Differentiate Minor Difference of Protein Structure in Biosimilar and Reference Products Using High-Resolution Orbitrap LC-MS/MS
Poster Note Disulfide Bond Analysis on Orbitrap Mass Spectrometry
Poster Note Rapid Peptide Mapping via Automated Integration of On-line Digestion, Separation and Mass Spectrometry for the Analysis of Therapeutic Proteins
Poster Note Complete Characterization of a Cysteine linked Antibody-Drug Conjugate Performed on a Hybrid Quadrupole-Orbitrap Mass Spectrometer with High Mass Range
Poster Note Orbitrap Based Mass Spectrometric Characterization of Antibody Drug Conjugates Engineered through Antibody Glycans
Poster Note Complete Characterization of a Cysteinelinked Antibody-Drug Conjugate Performed on a Hybrid Quadrupole-Orbitrap Mass Spectrometer with High Mass Range
Application Note Robust and reproducible peptide mapping and intact mass analysis workflows on a single instrument platform
Poster Note Characterization of Monoclonal Antibodies and ADCs using a Benchtop Orbitrap Mass Spectrometer
Poster Note Full Characterization of Heterogeneous Antibody Samples Under Denaturing and Native Conditions on a Hybrid Quadrupole-Orbitrap Mass Spectrometer
Poster Video Convenient High pH Peptide Fractionation
Poster Video High throughput Peptide Mapping by UHPLC-MS
Poster Video Improved Protein Identification using SubCellular Localization
Poster Video Orbitrap Based Mass Spectrometric Antibody Drug Conjugate Characterization
Presentation New, rapid and reproducible peptide mapping workflows
Product Spotlight SMART Digest Kit Q&A
Product Spotlight SMART Digest Soluble Trypsin Kit Smarter protein digestion
Technical Note High Resolution Peptide Mapping for Biopharmaceutical Analysis
Web Page Built For Biopharma Hypersite - Peptide Mapping
Web Page Peptide Mapping Webpage
Webinar Improved Characterization of Biotherapeutics with Better Peptide Mapping
Web Page BioPharma Finder Software - Planet Orbitrap
Web Page BioPharma Finder Software - Product page

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