Innovation for Biotherapeutic Peptide Mapping

Significance of the Topic

Comprehensive peptide mapping is fundamental for the characterization and quality control of biotherapeutics. It ensures accurate sequence verification, post-translational modification profiling and detection of low-abundance variants. Robust peptide mapping workflows accelerate development timelines, improve safety and support regulatory compliance in biopharma production.

Objectives and Study Overview

This whitepaper presents simplified, user-friendly peptide mapping solutions based on SCIEX mass spectrometry platforms and software. The goal is to demonstrate rapid method setup, comprehensive data acquisition and automated processing that enable both expert and novice users to obtain high-confidence peptide maps for monoclonal antibodies and other biologics.

Instrumentation Used

• X500B QTOF with SCIEX OS software
• TripleTOF 6600 high-resolution accurate-mass system
• BioPharmaView™ software for peptide data processing
• CESI 8000 Plus separation-ESI module for capillary electrophoresis

Methodology

Two data-acquisition strategies are compared: traditional information-dependent acquisition (IDA) and SWATH® acquisition. SWATH acquires high-resolution MS/MS spectra for all precursors, capturing low-abundance peptides and PTMs without pre-defined triggers. Workflows are configured using a point-and-click interface. BioPharmaView automates peptide identification, quantitation of modifications, glycopeptide profiling, deamidation mapping and disulfide bond localization.

Main Results and Discussion

• SWATH acquisition delivers unbiased, comprehensive peptide coverage (100% sequence mapping) in a single run.
• Low-abundance peptides and critical PTMs (oxidation, deamidation, glycosylation) are reliably detected and quantified.
• Automated glycopeptide detection enables confident assignment of G0, G1 and high-mannose species with MS/MS confirmation.
• Deamidated peptide sites are localized and relative ratios calculated easily through integrated visualization.
• Disulfide bond mapping yields high-resolution MS/MS spectra with clear annotation of multiple charge states.

Benefits and Practical Applications

• Rapid method development using SCIEX OS point-and-click setup reduces hands-on time.
• Automated data processing accelerates decision making and supports batch-to-batch comparability.
• Workflow simplicity makes peptide mapping accessible to non-specialists, broadening lab capability.
• Integration of orthogonal separation (CESI-MS) extends coverage for challenging peptides.

Future Trends and Opportunities

Advances in mass spectrometry are moving toward greater automation, machine-learning-driven data interpretation and cloud-based workflow management. Integration of multi-omic analyses, real-time process monitoring and high-throughput platforms will further streamline biologics characterization, enabling faster go/no-go decisions in early discovery and late-stage development.

Conclusion

The combined use of SCIEX high-resolution QTOF and TripleTOF instruments, SWATH acquisition and automated BioPharmaView processing offers a robust, user-friendly peptide mapping solution. This approach delivers complete sequence coverage, reliable PTM detection and simplified workflows, supporting efficient biotherapeutic development and quality control.