An Optimized SP3 Sample Processing Workflow for In-Depth and Reproducible Phosphoproteomics.

ABSTRACT: Protein phosphorylation is a ubiquitous post-translational modification (PTM) found across the kingdoms of life and is critical for the regulation of protein function in health and disease. Advances in high-throughput mass spectrometry have transformed our ability to interrogate the phosphoproteome. However, sample preparation methodologies optimized for phosphoproteomics have not kept pace, compromising the ability to fully exploit these technological advances. In this study, we present an optimized phosphoproteomics workflow using carboxylated SP3 magnetic beads, which have simplified proteomics sample preparation. By employing a washing step with 8 M urea and omitting the conventional C₁₈ SPE cleanup, we demonstrate a significant improvement in phosphopeptide identifications,

SUBMITTER: Daly LA 

PROVIDER: S-EPMC12322947 | biostudies-literature | 2025 Aug

REPOSITORIES: biostudies-literature