<p>Thiocyanate (SCN⁻) is an anionic contaminant and is a biomarker of environmental cyanide exposure. Therefore, it is particularly necessary to develop a simple, fast and sensitive method for SCN⁻ assay. Herein, a fluorescent assay method was developed to determine SCN⁻ based on Cu²⁺-triggered oxidation of o-phenylenediamine (OPD). Without SCN⁻, OPD was oxidized by Cu²⁺ and the resultant product (i.e., OPD oxide) exhibited a high fluorescence. However, in the presence of SCN⁻, Cu²⁺ reacted with SCN⁻; and thus, the oxidation ability of Cu²⁺ toward OPD was reduced, resulting in a weak fluorescence. Several important parameters such as the concentrations of Cu²⁺ and OPD, pH and the reaction time were optimized. Under the optimized conditions, the linear range was 5–80&#xa0;μM and the limit of detection was 5&#xa0;μM for SCN⁻ assay. In addition, the high selectivity and the satisfactory recoveries in tap water and artificial saliva were achieved for SCN⁻ detection due to the strong chelation between SCN⁻ and Cu²⁺. The whole detection process was simple and fast without using complex nanoparticles. More importantly, the developed assay method will have promising applications in monitoring of SCN⁻ for the assessment of water safety and human health.</p> Graphical abstract <p></p>