Quantitative plasma proteomics on stellar MS enables discovery of clinically relevant cancer biomarkers

Quantitative plasma proteomics is essential for discovering circulating biomarkers that enable early cancer detection and patient stratification. However, existing assays often lack the scalability, reproducibility, and sensitivity required for large clinical studies. To overcome these limitations, we developed a high-throughput targeted proteomics workflow on the Thermo Scientific™ Stellar™ mass spectrometer, enabling rapid and precise quantification of plasma proteins. The platform's fast acquisition speed and robust retention time stability allow comprehensive analysis of hundreds of peptides within a 30-minute gradient, while MS³ fragmentation enhances selectivity for low-abundance targets. Using the Biognosys PQ500 reference peptide kit, we quantified 804 peptides representing 322 plasma proteins, including 57 FDA-approved biomarkers, in plasma from patients with colorectal and lung cancer and healthy donors. More than 94% of peptides showed coefficients of variation below 25%, with linear responses across six orders of magnitude and limits of quantitation down to the attomole range. In colorectal cancer plasma, 29 proteins were significantly altered (adjusted p < 0.05, > 2-fold change) compared with controls. Notably, serum amyloid A2 (SAA2), alpha-2-glycoprotein-like (A2GL), and complement component C9 (CO9) were elevated-proteins implicated in inflammatory and immune pathways driving tumor progression. This study demonstrates that quantitative plasma proteomics on Stellar MS provides the sensitivity, reproducibility, and throughput needed to identify clinically relevant biomarker signatures in cancer. The workflow offers a robust and scalable foundation for translational proteomics and precision oncology applications, bridging the gap between discovery and clinical validation.