For the indirect determination of trace levels of organophosphorus pesticides in the environment, the irreversible inhibition of the activity of cholinesterases can be utilized with suitably designed electrochemical biosensors. This article reviews potentiometric and amperometric biosensors. For potentiometric biosensors, cholinesterases are immobilized on the active surface of pH-sensing electrodes, since during the biocatalytic reaction protons are produced as well as choline. In amperometric biosensors, choline oxidase is employed, in addition to esterases, which allows the indirect monitoring of choline by measurement of the consumption of oxygen or the production of hydrogen peroxide. The applications of various types of biosensor to the determination of pesticide residues in environmental and food analysis are also discussed.