Single-section integration of MALDI-MSI and Xenium spatial transcriptomics at single-cell resolution (nature.com)

• MALDI-MSI at 5 µm pixel size on Xenium slides, then same section processed for Xenium spatial transcriptomics.
• Transcript counts decreased by ~30% after MALDI-MSI, but cell recovery and cell-type assignments preserved.
• Custom software ESCDAT enables spatial alignment using MALDI laser ablation fiducials and cell boundary extraction.
• Integrated clustering in Seurat v5 shows enhanced cell-type resolution and reveals metabolic heterogeneity within RNA-defined populations.
• Demonstrated on mouse brain (validation) and human glioblastoma (proof-of-concept).

"This study presents a workflow combining high-resolution MALDI mass spectrometry imaging with Xenium spatial transcriptomics on the same tissue section, enabling pixel-scale co-registration of metabolic and transcriptomic data at single-cell resolution. Validated on mouse brain and demonstrated on human glioblastoma, the method preserves cell segmentation and cell-type assignment despite a ~30% reduction in transcript counts after MALDI imaging. The integration reveals metabolic heterogeneity within transcriptionally defined cell populations and allows per-cell matching of mass spectra to gene expression. This approach provides a scalable path for building multi-omic atlases."