<p>Heavy metal ions pollution in environmental waters has an increasing impact on human health. As two common metal ions, copper ions (Cu²⁺) and ferric ions (Fe³⁺) widely exist in nature and play a vital role in life process. Therefore, it is significant to design sensitive and simple detection approaches for Cu²⁺ and Fe³⁺. In our work, the ratiometric fluorescence analysis method (denoted as N-CDs/OPD) was established for Cu²⁺ and Fe³⁺ detection. The N-CDs exhibited a Cu²⁺ and Fe³⁺ fluorescence quenching response properties. The o-phenylenediamine (OPD) may be oxidized to 2,3-diaminophenazine (DAP) by Cu²⁺ and Fe³⁺. With addition of Cu²⁺ or Fe³⁺, the fluorescence of N-CDs (436&#xa0;nm) was quenched and a new peak at 556&#xa0;nm (DAP) appeared, which realized fluorescent ratiometric detection of Cu²⁺ and Fe³⁺. The Cu²⁺ concentration shows a good linear correlation versus fluorescence ratio (F₄₃₆/F₅₅₆) in the range of 10 to 30 µM (R² = 0.9981) with detection limit (LOD) of 0.86 µM. In addition, a good linear relationship between fluorescence ratio (F₄₃₆/F₅₅₆) and Fe³⁺ concentration in the range of 20 to 80 µM (R² = 0.9880) with LOD of 7.12 µM. This nanoprobe realizes the detection of authentic samples successfully, which is expected to serve as a testing kit for analysis in water samples.</p>