The binding properties of 18 monoclonal antibodies (MAbs) directed against the tobacco mosaic virus (TMV) coat protein were studied with five related tobamoviruses and seven mutant viruses, as well as with the dissociated coat proteins of these variants. Ten of the antibodies bound to both TMV and TMV protein, but these were able to discriminate between different mutants only when whole virus particles were compared in the immunoassay. Three MAbs reacted with TMV but not with dissociated viral subunits and these recognized the same residue exchanges. Five MAbs recognized synthetic peptides of 13-28 residues corresponding to parts of the protein. By comparing the common accessible surface between TMV and antigenically related viruses, it was possible to narrow down the region recognized by some of these MAbs. The linear peptide sequence recognized by these antibodies did not represent the entire epitope and residue exchanges around this region were able to affect the binding of MAbs.