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Protein engineering allows the generation of hybrid polypeptides with functional domains from di¡erent origins and therefore exhibiting new biological properties. We have explored several permissive sites in Escherichia coli L L-galactosidase to generate functional hybrid enzymes displaying a mouse scFv antibody fragment. When this segment was placed at the amino-terminus of the enzyme, the whole fusion protein was stable, maintained its speci¢c activity and interacted speci¢cally with the target antigen, a main antigenic determinant of foot-andmouth disease virus. In addition, the antigen-targeted enzyme was enzymatically active when bound to the antigen and therefore useful as a reagent in single-step immunoassays. These results prove the £exibility of E. coli L L-galactosidase as a carrier for large-sized functional domains with binding properties and prompt the further exploration of the biotechnological applicability of the scFv enzyme targeting principle for diagnosis or other biomedical applications involving antigen tagging.