Abstract: In order to enhance the sensitivity of acetylcholinesterase biosensor and detect organophosphorous pesticides effectively and rapidly, four different sources of enzymes used for detecting pesticides were tested and compared. Homogenates from chicken liver, chicken brain and cyprinoids brain, respectively were home-made in laboratory and acetylcholinesterase (AChE, from Musca domestica) was purchased. The inhibition rates of AChE activities under different concentrations of omethoate, trichlorfon and dichlorvos were compared. The results showed that the AChE activities in chicken liver and cyprinoids brain were very low. The capacity of activity inhibition of chicken brain AChE was the best in four kinds of enzymes. Inhibition rates of chicken brain-esterase achieved 24.16%, 50.56%, 54.1% for 100μg/L of omethoate, trichlorfon and dichlorvos, respectively. Chicken brain-esterase had a high specific activity, and it was cheap and easy to be purified, and the inhibition sensitivity of chicken brain-esterase to three organophosphorus pesticides was significantly higher than that of chicken liver and cyprinoids brain. So the chicken brain-esterase can be used to detect pesticide residues in agricultural products.