In this paper, a novel turn-on sensor was constructed for quantification and imaging of acetamiprid. Acetamiprid aptamer-modified ZnS:Mn probe (ZnS:Mn-Aptamer) was obtained by conjugating ZnS:Mn QDs with acetamiprid binding aptamer. The fluorescence of the probe was turn off by multi-walled carbon nanotubes (MWCNTs) based on fluorescence resonance energy transfer (FRET) between ZnS:Mn-Aptamer and MWCNTs. After acetamiprid was introduced, the fluorescence was turn on because ZnS:Mn-Aptamer specifically binded with acetamiprid. Based on the above-mentioned principle, the quantitative detection and imaging of acetamiprid in real samples were achieved. The linear equation could be described as F/F0=1.101+0.0312CA (CA represented the concentration of acetamiprid, nM) in a linear range of 0–150nM with a detection limit of 0.7nM. The results suggested this method had remarkable sensitivity, selectivity, and was very simple without complex operation. Meanwhile, this novel turn-on sensor has the potential of in situ visual determination for pesticide residues in complex system.