Discriminating Live and Dead Cells in Dual-Color Mode with a Two-Photon Fluorescent Probe Based on ESIPT Mechanism

Discrimination of live and dead cells is an important task in biological, pathological, medical, and pharmaceutical studies. In this work, we have developed a novel fluorescent probe DACA that can discriminate live and dead cells in a dual-color mode for the first time. DACA can stain dead cells with blue fluorescence peaked at 440 nm, while it can also label live cells with orange emission peaked at 570 nm. Compared with one-color fluorescent probes, such a dual-color probe can efficiently avoid false positive results from cellular autofluorescence and misleading signals brought by inhomogeneous staining, and thus can supply more accurate information in biological applications. By means of DACA, the health status of tumor cells pretreated by H₂O₂ and ultraviolet radiation has been successfully detected and imaged. Moreover, DACA and the hydrolyzed product exhibit excellent two-photon properties. Live and dead cells, as well as the zebrafishes, have been discriminated with dual emission colors under one- and two-photon microscope. These results demonstrate that DACA is a powerful tool for dual-color distinguishing live and dead cells in vitro and in vivo.