In this study, we have constructed a simple, sensitive and rapid biosensor for detection of acetylcholine esterase (AChE) based on CdTe quantum dots (QDs). The detection limit of AChE by one-step enzyme reaction based on the thiolglycolic acid (TGA) stabilized QDs (TGA-QDs) was 10 U/L and the linear range was 10-100 and 100-1200 U/L, respectively. The detection limit of AChE by two-step enzyme reaction based on the 3-mercaptopropionic acid (MGA) stabilized QDs (MGA-QDs) was found to be 20 U/L and the linear range was 100-2500 U/L. The experimental conditions of biosensors were optimized, and the detection mechanism was studied. We also detected AChE in serum samples based on TGA-QDs or MGA-QDs. The linear range was 10-140 and 50-1000 U/L, respectively. The excellent performance of this novel biosensor demonstrated that this strategy has prodigious potential to be applied in practice detection of AChE. The detection limit of AChE by one-step enzyme reaction based on the thiolglycolic acid (TGA). This detection limit of AChE by one-step enzyme reaction based on the thiolglycolic acid (TGA) The detection limit of AChE by two-step enzyme reaction based on the 3-mercaptopropionic acid (MGA (TGA-QDs) was 10 U / L and the linear range was 10-100 and 100-1200 U / L, ) stabilized QDs (MGA-QDs) was found to be 20 U / L and the linear range was 100-2500 U / L. The experimental conditions of biosensors were optimized, and the detection mechanism was studied. We also detected AChE in serum samples based on TGA-QDs or MGA-QDs. The linear range was 10-140 and 50-1000 U / L, respectively. The excellent performance of this novel biosensor for that strategy has prodigious potential to be applied in practice detection of AChE.