A novel disposable electrochemical immunosensor for highly selective and sensitive detection of organophosphorylated butyrylcholinesterase (OP‐BChE), a specific biomarker for exposure to toxic organophosphorus agents, is presented. In this new approach, zirconia nanoparticles (ZrO2) were employed to selectively capture the OP moiety of OP‐BChE adducts, followed by quantum dot (QD)‐tagged anti‐BChE antibodies for amplified quantification. The captured CdSe‐QD tags can be sensitively detected by stripping voltammetry using an in situ bismuth‐plating method. The OP agent, diisopropylfluorophosphate (DFP), was selected to prepare OP‐BChE adducts in various matrices. The formation of OP‐BChE adducts in plasma sample was confirmed using mass spectroscopy. The developed electrochemical immunosensor demonstrates a highly linear voltammetric response over the range of 0.1 to 30 nM OP‐BChE, with a detection limit of 0.03 nM (based on signal/noise = 3), coupled with a good reproducibility (relative standard deviation 4.5%). Moreover, the immunosensor has been validated with biomonitoring of OP‐BChE adducts in the plasma samples. This novel nanoparticle‐based electrochemical immunosensor thus provides an alternative way for designing a sensitive and cost‐effective sensing platform for on‐site screening/evaluating exposure to a variety of OP agents.