Cyprinid herpesvirus 3 (CyHV-3) is the pathogenic agent of koi herpesvirus disease (KHVD) afflicting common carp and koi (Cyprinus carpio L.) populations globally. As described previously, proteomic analyses of purified CyHV-3 particles have shown that at least 46 structural proteins are incorporated into CyHV-3 virions; among these ORF136 may encode a putative envelope protein. In this study, Western blotting analysis showed that a specific band with the predicted molecular weight of 17 kDa was detected both in purified virions and envelope components using a rabbit anti-ORF136 polyclonal antibody. Indirect immunofluorescence assay with confocal laser scanning microscopy indicated that the ORF136 protein was distributed in the cytoplasm of CCB cells infected with CyHV-3 and transfected with a pVAX1-ORF136 plasmid. Furthermore, immunogold electron microscopy confirmed that ORF136 protein localized to the CyHV-3 envelope.