Ratiometric fluorescent sensors for detecting zinc ions in aqueous solution and living cells with two-photon microscopy

We report two fluorescent Zn²⁺ sensors, AD1 and AD2. In aqueous solution, upon addition of Zn²⁺, AD1 only showed the enhancement on the fluorescence intensity, while AD2, acting as a ratiometric sensor, demonstrated the variations not only on the fluorescence intensity but also on the emission wavelengths. Both sensors exhibit high sensitivity (nM level) and selectivity for Zn²⁺ over Na⁺, K⁺, Mg²⁺, and Ca²⁺ at the millimolar level. Moreover, AD2 undergoes significant increase of the TPA cross sections (δ_max) at 840 nm by the Zn²⁺-coordination induced ICT enhancement. The laser scanning confocal microscopy experiments revealed that AD2 is cell-permeable and can indeed visualize the changes of intracellular Zn²⁺ in living cells through a ratiometric approach by utilizing two-photon excitation.