Mass spectrometry-based human spatial omics: fundamentals, innovations, and applications

Abstract Mass spectrometry-based spatial omics is a powerful approach for visualizing the spatial organization of proteins, metabolites, lipids, and other biomolecules in situ, combining the molecular depth of mass spectrometry with spatially resolved imaging. This systematic review traces the rapid technological and computational evolution of this field, including innovations in mass spectrometry imaging (MSI), labeling-based approaches, and proximity labeling techniques. It also highlights recent advances that enhance spatial resolution, expand molecular coverage, and enable deep molecular characterization and review analytical pipelines that integrate deep learning, cross-modality registration, and cloud-optimized data formats. From the multimodal and practical perspective, the integration of MSI with other spatial omics platforms and its transformative applications in tumor microenvironment profiling, neurodegenerative disease, developmental biology, biomarker discovery, and precision medicine are discussed. Finally, this review outlines challenges and opportunities, emphasizing the need for standardization, clinical validation, and interpretable artificial intelligence to enable broader adoption. These advances position MS-based spatial omics as a foundational pillar for multimodal spatial biology and personalized healthcare.