Ultrafast deep-red emission fluorescent probe for highly selective imaging of endogenous cysteine in living cells and mice

It is still challenging how to selectively and sensitively detect cysteine in living systems which plays crucial roles in many physiological and biological processes. Here, we developed a deep-red emission fluorescent probe (DRP-Cys) for Cys based on a hemicyanine scaffold. A tailed malononitrile group could chromatically shift excitation/emission wavelengths to avoid the limitations such as photo-damage or auto-fluorescence in biological samples. Compared with other amino acids, Cys could trigger the Michael addition reaction to release a dramatic enhancement in fluorescence centering at around 645 nm in DMSO aqueous solution under physiological conditions with an ultrafast response time of about 1 min and a low limit of detection of 10 nM which significantly benefited from the introduction of the malononitrile moiety. The probe was also successfully applied for visualizing endogenous Cys in living HeLa cells and mice with low cytotoxicity and good cell-membrane permeability.