A water-soluble, cell-permeable Mn(ii) sensor enables visualization of manganese dynamics in live mammalian cells.

Central roles of Mn(2+) ions in immunity, brain function, and photosynthesis necessitate probes for tracking this essential metal ion in living systems. However, developing a cell-permeable, fluorescent sensor for selective imaging of Mn(2+) ions in the aqueous cellular milieu has remained a challenge. This is because Mn(2+) is a weak binder to ligand-scaffolds and Mn(2+) ions quench fluorescent dyes leading to turn-off sensors that are not applicable for in vivo imaging. Sensors with a unique combination of Mn(2+) selectivity, μM sensitivity, and response in aqueous media are necessary for not only visualizing labile cellular Mn(2+) ions live, but also for measuring Mn(2+) concentrations in living cells. No sensor has achieved this combination thus far. Here we report a novel, completely water-soluble, reversible, fluorescent turn-on, Mn(2+) selective sensor, M4, with a K (d) of 1.4 μM for Mn(2+) ions. M4 entered cells within 15 min of direct incubation and was applied to image Mn(2+) ions in living mammalian cells in both confocal fluorescence intensity and lifetime-based set-ups. The probe was able to visualize Mn(2+) dynamics in live cells revealing differential Mn(2+) localization and uptake dynamics under pathophysiological versus physiological conditions. In a key experiment, we generated an in-cell Mn(2+) response curve for the sensor which allowed the measurement of the endogenous labile Mn(2+) concentration in HeLa cells as 1.14 ± 0.15 μM. Thus, our computationally designed, selective, sensitive, and cell-permeable sensor with a 620 nM limit of detection for Mn(2+) in water provides the first estimate of endogenous labile Mn(2+) levels in mammalian cells.