Baculovirus-expressed foot-and-mouth disease virus (FMDV) nonstructural proteins 2C and 3D were used as the antigens in a western blotting assay. This assay allowed for differentiation of FMDV-infected pigs from vaccinated pigs. Serial studies were performed using sera collected from experimentally infected and vaccinated pigs. Positive reactions were found in the sera derived from the experimentally infected pigs. There was, however, no positive reaction in the vaccinated pigs. These results suggest that this western blotting assay is a useful method for differentiation of infected pigs from vaccinated pigs.