Abstract Regions rich in 5-methylcytosine were localized in male metaphase chromosomes of the Indian muntjac deer ( Muntiakus muntjak). Chromosomes were ultraviolet irradiated and subsequently photooxidized in the presence of methylene blue to induce maximum DNA denaturation. Following treatment with anti 5-methylcytosine antibody (anti 5-MeC), regions of antibody binding were visualized by an immunofluorescence or immunopreoxidase staining procedure. All chromosomes showed some level of antibody binding along their length and at centromeric regions, with intense binding evident in the centromere of chromosome 3 and the elongated centromeric “neck” of chromosome 3-X. The Y chromosome displayed low levels of antibody binding. The banding pattern observed with anti 5-MeC is the reverse of that obtained by quinacrine staining.