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Enzyme-linked immunosorbent assays (ELISAs) for the class-specific determination of organophosphorus (OP) pesticides were developed from monoclonal antibodies raised against haptens with the functional group common to OP pesticides. To develop antigen-coated, indirect, competitive ELISAs, four haptens with different spacer arm structures were used to prepare antibodies, while eight haptens were tested for use as coating antigens. A total of 32 ELISAs were developed with one selected as the most suitable one based on average IC(50) and % CV values. The chosen ELISA showed class-selective response to O,O-diethyl phosphorothioate and phosphorodithioate OP pesticides with negligible cross-reactivity to other types of pesticides. Average IC(50) and % CV values of this ELISA for the 12 OP pesticides were 89 ng/mL and 96%, respectively. Compared to ELISAs previously developed with the same objective, the current ELISA demonstrates better sensitivity based on much lower mean IC(50) values in addition to improved class-selective determination based on considerably lower % CV values as well as precise discrimination against other types of pesticides.