Singlet oxygen (O-1(2)) is an important species for oxidation in biological processes. O-1(2) is implicated in the genotoxic effect, and plays an important role in the cell-signaling cascade and in the induction of gene expression. However, the rapid detection of O-1(2) in biological environments with sufficient specificity and sensitivity is hampered by its extremely low emission probability. Here, a layer-by-layer (LbL) film of CdTe quantum dots (QDs), polymers, and ascorbate have been designed as a rapid, highly selective, and sensitive fluorescence probe for O-1(2) detection. Upon reaction with O-1(2), the probe exhibits a strong photoluminescence (PL) response even at trace levels. This remarkable PL change should enable the probe to be used for O-1(2) detection in many chemical and biological systems and as an environmental sensor.