Enzyme‐based electrochemical biosensors with sufficient sensing specificity are useful analytical tools for detection of biologically important substances in complicated systems. Here, we present the design of a nano‐hybrid biosensor for the specific and sensitive detection of methyl parathion (MP). The nano‐hybrid sensing film was prepared via the formation of Au nanoparticals (AuNPs) on silica nanoparticles (SiNPs), mixing with multiwall carbon nanotube (MWNTs) and subsequent immobilization of methyl parathion degrading enzyme (MPD). The fabrication procedure was characterized by scanning electron images, linear scan voltammetry and electrochemical impedance spectroscopy. The combined MPD exhibited high affinity to it substrate and thus a selective, sensitive, fast and cheap method for determination of MP, quantitatively was proposed. A significant synergistic effect of nano‐hybrid on the biosensor performance was observed in biosensing MP. The square wave voltammetric responses displayed well defined peaks, linearly proportional to the concentration of MP in the range from 0.001 to 5.0 μg/mL with a detection limit of 0.3 ng/mL. The proposed biosensor also showed good precision and reproducibility, acceptable stability and accuracy in garlic samples analysis. It provided a platform for the simple and fast construction of biosensors with good performance for the determination of enzyme‐specific electroactive species.