NZW rabbits (,2 kg body weight) were purchased from Millbrook Breeding Labs (Amherst, MA) for immunogenicity studies, and housed in the Department of Animal Medicine at the University of Massachusetts Medical School in accordance with IACUC approved protocol. The rabbits (3 rabbits/group) were immunized with a Helios gene gun (Bio-Rad) at the shaved abdominal skin as previously reported [44] with a total of 36 mg H5 HA DNA vaccine plasmid or vector control plasmid at each immunization. DNA immunizations were given at weeks 0, 2, 4, 8. Serum samples were taken prior to the first immunization and 2 weeks after each immunization for study of H5 HA-specific antibody responses.