Synthesis of Cyclopropene‐Modified Fatty Acids Allows Single‐Cell Quantification of Uptake by Immune Cells

Immune cell activity is strongly influenced by the nutrients present during activation. The effects of specific fatty acids (FAs) can be particularly complex, with them exerting diverse and sometimes opposing effects on immune cells. These functional differences are thought to stem from structural differences between the FAs, leading to altered cellular handling. However, chemical tools to directly probe these aspects remain limited. Here, we report the design and synthesis of saturated, unsaturated, and polyunsaturated cyclopropenyl fatty acids, each incorporating a minimal one-carbon cyclopropene moiety as a bioorthogonal click handle. This motif enables rapid and live-cell-compatible labeling via the inverse electron-demand Diels-Alder reaction, providing a versatile platform to trace FA behavior in biological systems. Application of these synthetic cyclopropenyl FAs in primary immune cell mixtures reveals distinct uptake patterns between different cell types, with polyunsaturated analogues showing strong uptake across all immune cell types. Complementary metabolic and proteomic analyses suggest that this uptake results in biological differences between high/low uptake immune populations, highlighting the utility of cyclopropenyl FA probes for dissecting lipid uptake in the context of immune cell biology.