To investigate the antigenic nature of cylindrical inclusion proteins (CIPs) of the potyviruses Turnip mosaic virus (TuMV) and Zucchini yellow mosaic virus (ZYMV), monoclonal antibodies (MAbs) against the two CIPs were produced and epitopes on the CIPs were localized using Escherichia coli-expressed CIP fragments in Western blot analysis. All 23 MAbs against ZYMV CIP reacted only with ZYMV CIP. In contrast out of the 18 MAbs produced against TuMV CIP, 14 MAbs were TuMV CIP-specific while the remaining four MAbs cross-reacted with both CIPs. The four cross-reactive MAbs recognized two distinct epitopes in the N-terminal half of TuMV CIP corresponding to amino acid residues 103-119 and 224-237. Thirteen out of 14 TuMV CIP-specific MAbs recognized two distinct epitopes within residues 1-102 and 120-214, while the other one recognized an epitope within residues 301-644. On the other hand, 21 out of 23 ZYMV CIP-specific MAbs recognized epitopes within residues 1-118, while the remaining two recognized epitopes within residues 301-522. These results suggest that cross-reactive and major virus-specific epitopes are located at the N-terminal half of the respective CIPs.