Accurate Measurement of Peptide Variant Mixtures with Next-Generation Protein Sequencing™ (NGPS™)

US HUPO 2025 Poster – ProteoVue™ is a comprehensive bioinformatics pipeline for single-amino acid variant (SAAV) detection and quantification using the Quantum-Si Platinum® and Platinum® Pro NGPS instruments. ProteoVue integrates pulse calling, recognition segment detection, fluorescence dye classification, and a neural network-driven kinetic signature database to enable variant calling with high sensitivity and specificity. These components feed into a scoring-based alignment and clustering framework, allowing accurate differentiation of binary peptide mixtures and proteoforms with subtle sequence variations. We demonstrate that ProteoVue successfully detects SAAVs across a range of substitution types, including variants lacking direct recognizer interactions, by leveraging kinetic features and sequence context. While some substitutions present challenges, the pipeline consistently captures key kinetic signatures for variant discrimination, underscoring its potential as a versatile tool for proteomic research. As NGPS technology matures and recognizer libraries expand, ProteoVue will enable more refined variant analysis, supporting applications in basic research, biomarker discovery, and clinical proteomics.