Raccoonpox virus (RCN) has been used as a recombinant vector against several mammalian pathogens but has not been tested in birds. The replication of RCN in chick embryo fibroblasts (CEFs) and chickens was studied with the use of highly pathogenic avian influenza virus H5N1 hemagglutinin (HA) as a model antigen and luciferase (luc) as a reporter gene. Although RCN replicated to low levels in CEFs, it efficiently expressed recombinant proteins and, in vivo, elicited anti-HA immunoglobulin yolk (IgY) antibody responses comparable to inactivated influenza virus. Biophotonic in vivo imaging of 1-wk-old chicks with RCN-luc showed strong expression of the luc reporter gene lasting up to 3 days postinfection. These studies demonstrate the potential of RCN as a vaccine vector for avian influenza and other poultry pathogens. ?? American Association of Avian Pathologists 2010.