Optimized MALDI2-Mass Spectrometry Imaging for Stable Isotope Tracing of Tissue-Specific Metabolic Pathways in Mice

-glucose across glycolysis, glycogen metabolism, and the tricarboxylic acid (TCA) cycle in kidney tissues. In brain sections, we successfully mapped six intermediates from the TCA cycle and glutamate-glutamine (Glu-Gln) cycle simultaneously in distinct neurological regions. Furthermore, in breast cancer tumor tissues, our approach facilitated the mapping of nine metabolic intermediates in multiple pathways, including glycolysis, the pentose phosphate pathway (PPP), and the TCA cycle, illustrating metabolic heterogeneity within the tumor microenvironment. This methodology enhances metabolite coverage, enabling more comprehensive imaging of isotope-labeled metabolites and opening new avenues for exploring the metabolic landscape in various biological contexts.