Highly pathogenic avian influenza H5N1 virus represents a growing threat for an influenza pandemic. Development of effective vaccines for H5N1 is a priority for pandemic preparedness. Focusing on influenza virus-like particles (VLPs) has been suggested as a promising vaccine approach. Recent VLP vaccination efforts have been concentrated on the H5N1 strains isolated from humans. Because all confirmed cases of human H5N1 infection were directly transmitted from infected poultry, it is of interest to develop VLP vaccines comprised of antigenic proteins of avian H5N1 strains in order to compare their efficacy in fighting diverse H5N1 strains with vaccines developed using human isolates. In this study, we generated a VLP vaccine composed of the HA, NA, and M1 proteins of the avian H5N1 influenza virus isolate A/chicken/Hubei/489/2004, which seems to occupy a unique phylogenetic position; it belongs to neither clade 1 nor clade 2. Upon infection of Sf9 insect cells using recombinant baculoviruses, the co-expressed HA, NA, and M1 proteins self-assembled and released into the culture medium as VLPs. In a mouse model, purified VLPs elicited an effective antibody response and conferred complete protection against heterologous human H5N1 influenza virus, as well as a homologous avian H5N1 influenza virus isolate. Our work provides further evidence that vaccination with influenza VLPs may be a productive approach to achieve protection against diverse H5N1 strains.