Cyprinid herpesvirus II (CyHV-2) is the main pathogen causing haematopoietic necrosis disease in . Although many nucleic acid-based diagnostic methods have been applied, no stable and sensitive immunological diagnostic methods have been reported. In this study, to detect CyHV-2 in clinical samples by immunological methods, recombinant protein 72, encoded by the CyHV-2 ORF72 gene, was used as a capture antigen to identify specific antibodiesagainst CyHV-2 in the serum of infected fish. First, ORF72 was amplified from the purified CyHV-2 genome, and cloned into expression vector PGEX-4t-3 to produce recombinant GST-ORF72 in . The recombinant protein was detected by sodium dodecyl sulphate-polyacrylamide gel electrophoresis. The purified recombinant protein 72 was used to immunize mice and polyclonal antibodies were obtained. Western blotting showed that the polyclonal antibody could not only identify the recombinant protein, but could also recognize protein 72 in virus particles. A Western blotting protocol was established for the specific detection of antibodies against protein 72 protein of CyHV-2 and then applied to detect CyHV-2 infection in clinical fish sera samples. In conclusion, the Western blotting technique established in this study could be used for specific serodiagnosis of CyHV-2 infection.