Automation streamlines peptide map preparation, analysis and reporting for biotherapeutic antibody characterization

Among various analytical methods for characterizing biotherapeutic antibodies, peptide mapping is one of the most widely used due to its unparalleled sensitivity and specificity. A typical peptide mapping workflow includes enzymatic digestion of a protein into peptides, peptide mixture separation and detection by liquid chromatography coupled with mass spectrometry, and data analysis. This analysis offers detailed structural insights into the protein sequence, post-translational modifications (PTM), and process-related impurities. Such information is essential for drug product and process development, including critical quality attribute (CQA) assessment, cell line selection, and production process optimization. Various peptide map methods are typically used for protein characterization. Besides the commonly used tryptic map for protein sequence coverage and PTM analysis, specialized peptide map methods include the Lys-C (lysyl-endopeptidase) map for disulfide linkage confirmation and cysteine modification identification, and the Asp-N map (using endoproteinase Asp-N) for site-specific glycation determination, among others. Protein digestion, data analysis, and reporting can significantly extend turnaround timelines. To address this challenge, automated protein digestion protocols using liquid handlers were implemented to replace lengthy manual sample preparations. Additionally, a streamlined, compliance-ready data analysis workflow for data analysis and reporting was developed. Automating these steps from sample preparation to data reporting has transformed time-consuming peptide mapping workflows into streamlined automated workflows. This significantly improved throughput, leading to 2-3 days saved per week (a 30 % improvement), enhanced accessibility, and overall user friendliness.