Abstract: The acetyl cholinesterase biosensors modified by carbon nanotubes were prepared by drop coating, self-assembly and chemical bonding methods. The electrochemical sensing properties and morphology of biosensor was characterized through cyclic voltammetry (CV), chronoamperometry (CA), electrochemical impedance spectroscopy (EIS) and scanning electron microscope (SEM). The performance of the biosensor analyzing typical carbamate pesticide isoprocarb was investigated. The kinetic properties and electrochemical behavior of each biosensor was researched. A new equivalent circuit model on immobilized enzyme biosensor was built based on these findings. The results showed that, compared to bare electrode, the apparent surface areas of each biosensor was substantial increased, and the electron transfer rate of each biosensor decreased in an order as follows: acetyl cholinesterase/chitosan/functionalized carbon nanotubes biosensor>acetylcholinesterase/functionalized carbon nanotubesbiosensor>chitosan/acetylcholinesterase/functionalized carbon nanotubes biosensor (AChE/CS/F-CNTs/GCE>AChE/F-CNTs/GCE>CS/DAC-AChE/F-CNTs/GCE). The constants of each biosensor apparent electron transfer rate were: ks AChE/CS/F-CNTs/GCE=0.24 s-1, ks AChE/F-CNTs/GCE=0.23 s-1, ks CS/DAC-AChE/F-CNTs/GCE=0.22 s-1. The equivalent circuit model of electrical impedance spectroscopy was R1(CPE1(R2(CPE2(R3)))) and the specific device parameters of each circuit model were figured out. The results confirmed that the equivalent circuit can effectively simulate the sensing mechanism of sensor detecting the isoprocarb. It would be a reference for the rapid detection of pesticide residue by biosensors.