Fluorescence polarization immunoassay for the determination of sulfamethazine was developed via four polyclonal antibodies against sufamethazine and five fluorescein-labeled sulfonamides(tracers). The study mainly focuses on the relative affinity trend of four polycolnal antibodies toward 17 sulfonamides analogs and explaination of the recognition between antibody and drugs based on the two-dimensional structures of drug antigen. The results demonstrate that the polyclonal antibody(No.3) showed the most sensitivity for sulfamethazine,however,the four polyclonal antibodies presented a similar relative affinity trend to 17 sulfonamides,implying that the affinity of antibody to antigen has correlationship with the chemical structure of antigen.