Based on conjugate addition-cyclization reaction of Cys with acrylate and subsequent 1,6-elimination of p-hydroxybenzyl moiety, a novel colorimetric and ratiometric fluorescent probe 1 was designed and synthesized. Upon addition of Cys to the solution of 1, the absorption spectra changed from 508 nm to 452 nm (Δ56 nm) and afforded visible color change from pink to yellow. Meanwhile, the emission spectra shifted from 644 nm to 539 nm (Δ105 nm) with remarkable changes in the emission ratio of F539 nm/F644 nm (R/R0 up to 760-fold), accompanying with an obvious fluorescence change from orange to green under illumination with a 365 nm UV lamp. In addition, 1 exhibited a large Stokes shift (136 nm), high sensitivity (detection limit of 46.7 nM), and excellent selectivity to Cys over Hcy and GSH. Moreover, 1 can discriminate Cys from Hcy and GSH by fluorescence spectra, even obvious visible and fluorescence color changes. Importantly, 1 can be used to image Cys in living cells by dual emission channels.