Epitopes of strain A22 Iraq 24/64 of foot-and-mouth disease virus have been mapped with monoclonal antibodies (MAbs). Three methods were used: (i) an indirect ELISA using an overlapping set of peptides, (ii) production of neutralization escape variants against each MAb and (iii) sequencing of neutralization escape variants. The study has shown that the virus has at least three overlapping liner neutralizing epitopes within a major antigenic site on VP1. The presence of a second, conformational site was demonstrated but its position on the virus particle was not located. Synthetic peptides with sequences representing the major site elicit antibodies which have similar broad cross-neutralizing activity to polyclonal serum or neutralizing MAbs produced with the virus against a range of field isolates.