A unique type of pyrrole-based cyanine fluorophores with turn-on and ratiometric fluorescence signals at different pH regions for sensing pH in enzymes and living cells.

The development of new functional fluorescent dyes has attracted great attention. Herein we have described a novel strategy to design a unique type of cyanine dyes by attaching two indolium moieties at the α-positions of the pyrrole core. The new type of cyanine dyes is named as PyCy fluorophores. Importantly, PyCy dyes can exhibit an exceptional feature, fluorescence turn-on response at pH varying from acidic to near-neutral conditions, and a ratiometric fluorescence response at pH varying from near-neutral to basic conditions. By taking advantage of the fluorescence turn-on response of PyCy2 at pH varying from acidic to near-neutral conditions and emission properties of PyCy2, we have demonstrated that a small-molecule fluorescent probe can image pH variations in living cells. Furthermore, we have demonstrated that PyCy2 can sense real-time pH changes under alkaline conditions induced by enzymes based on the ratiometric fluorescence response of PyCy2 at pH varying from near-neutral to basic conditions. We expect that the new design strategy for PyCy fluorophores may prompt the development of a wide variety of cyanine derivatives with desirable properties.