Liquid and gas chromatography are commonly used to measure organophosphorus pesticides. However, these methods are relatively time consuming and require a tedious sample pretreatment. Here, we applied the localized surface plasmon resonance (LSPR) of gold nanoparticles covalently coupled with acetylcholinesterase (AChE) to create a biosensor for detecting an example of serial signals responding to paraoxon in the range of 1-100 ppb by an AChE modified LSPR sensor immersing in a 0.05 mM ACh solution. The underlying mechanism is that paraoxon prevents acetylcholine chloride (ACh) reacting with AChE by destroying the OH bond of serine in AChE. We found that the AChE modified LSPR sensors prepared by incubation with 12.5 mU/mL of AChE in phosphate buffer solution at pH 8.5 room temperature for 14 h have the best linear inhibition response with a 0.234 ppb limit of paraoxon detection. A 14% of inhibition on the sensor corresponds to the change of paraoxon concentration from 1 to 100 ppb. The sensor remained 94% of its original activity after six cycles of inhibition with 500 ppb paraoxon followed with reactivation of AChE by 0.5 mM 2-pyriding-aldoxime methoiodide (2-PAM). In addition, the sensor retains activity and gives reproducible results after storage in dry state at 4 degrees C for 60 days. In conclusion, we demonstrated that the AChE modified LSPR sensors can be used to determine the concentration of paraoxon biosensor with high sensitive and stable characteristics.