Functions of antibody are determined by its structure. However, the interaction mechanism is still not known since no carbaryl antibody crystallization was analyzed, which severely affects its application in ELISA detection. This study aimed to predict the 3D structure and binding mode of anti-carbaryl scFv with antigen. The molecular modeling and docking technology were employed by utilizing relative bioinformatics tools and on-line resources. The results showed a hydrophobic groove was formed by H-CDR2, H-CDR3 and L-CDR2, which block in carbaryl firmly. The binding site was composed of Ala51, Ser52, Ile51, Gly54, Ser56, Arg98 and Gly100, etc. These data highlighted the mechanism of interaction between anti-carbaryl antibody and antigen. Furthermore, it provides guidance for in vitro affinity maturation of anti-carbaryl antibody.