Tensile Expansion Mass Spectrometry for single cell metabolomics imaging

Abstract Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) enables the spatial mapping of endogenous biomolecules within native biological specimens; however, it remains limited in achieving single-cell resolution. While advances in instrument modifications, computational processing methods, and tissue-based sample preparation have facilitated high lateral resolutions and cellular level imaging, resolving metabolic heterogeneity at the single-cell level remains challenging for users without specific expertise or custom instrumentation. Here, we present tensile expansion mass spectrometry (TExMS), a cost-effective approach for single-cell MALDI-MSI that is compatible with commercial MSI instrumentation. TExMS utilizes highly stretchable hydrogels as a substrate for live-cell seeding, attachment, and desiccation, avoiding the need for chemical fixation and enabling the retention of both intracellular and extracellular metabolites, including media-derived components that are lost during fixation and washing. We used TExMS to expand individual cells of a human high-grade serous ovarian cancer (HGSOC) cell line and spatially map their small molecule (