Cellular metabolic plasticity enables cells to meet rapidly changing biosynthetic demands in both physiology and disease. Yet interrogating this complexity at subcellular resolution has been limited by existing metabolic imaging approaches, which typically track only a single pathway at a time. As a result, it has been difficult to obtain a systems-level view of coordinated precursor utilization and turnover across interconnected metabolic networks.
To address this gap, we developed SuMMIT-SRS (Subcellular Multiplexed Metabolic Isotope Tracing via Stimulated Raman Scattering microscopy), a platform that enables simultaneous, subcellular-resolution visualization of DNA, RNA, protein, and lipid metabolic dynamics. SuMMIT-SRS leverages the distinct vibrational signatures of carbon–deuterium (C–D) bonds introduced through a palette of deuterated precursors—including amino acids, lipids, and monosaccharides—to map co-regulated biosynthetic programs and disentangle amino acid–specific metabolic pathways in intact cells and tissues.
We demonstrate the broad applicability of SuMMIT-SRS by capturing cell type–specific shifts in substrate usage across diverse models, including breast cancer organoids, the Drosophila fat body and developing brain, aged human neurons, and mouse liver under genetic and pathological perturbations. By extending SRS microscopy to multiplexed isotope tracing, SuMMIT-SRS provides a powerful approach for revealing dynamic biosynthetic programs in development, homeostasis, aging, and disease.
