A simple and effective strategy for designing ratiometric fluorescent nanosensor has been described in this work. A carbon dots (CDs) based dual-emission nanosensor for Cu(2+) detection was prepared by coating CDs on the surface of Rhodamine B-doped silica nanoparticles. The fluorescent CDs were synthesized using N-(β-aminoethyl)-γ-aminopropyl methyldimethoxysilane (AEAPMS) as the main raw material, so that the residual ethylenediamine groups and methoxysilane groups on the surface of CDs can serve as the Cu(2+) recognition sites and the silylation reaction groups. The obtained nanosensor showed characteristic fluorescence emissions of Rhodamine B (red) and CDs (blue) under a single excitation wavelength. Upon binding to Cu(2+), only the fluorescence of CDs was quenched, resulting in the ratiometric fluorescence response of the dual-emission silica nanoparticles. This ratiometric nanosensor exhibited good selectivity to Cu(2+) over other substances, such as metal ions, amino acids, proteins, and vitamin C. The ratio of F467/F585 linearly decreased with the increasing of Cu(2+) concentration in the range of 0 to 3 × 10(-6) M, a detection limit as low as 35.2 nM was achieved. Additionally, this nanosensor was successfully applied for the ratiometric fluorescence imaging of Cu(2+) in cells and determination of Cu(2+) in real tap water.