To obtain AdHu5-HA vaccine, HEK 293 cells were transfected with 10 mg of linearized pAdenoX-HA DNA in calcium phosphate (BD Biosciences) solution and cells were cultured until the appearance of cytopathic effects (CPE). Similarly, VR1BL E1 cells were transfected with 10 ug of linearized pFPAV227-HA DNA combined with Lipofectin (Invitrogen) and cells were cultured until CPE were apparent. Amplified adenoviruses containing cell lysates were harvested, freeze-thawed three times, and purified by CsCl gradients. The integrity of the H05-HA transgene cassette was confirmed through EcoRI restriction digests and by sequencing (DNA Core, National Microbiology Laboratory) with multiple primer sets. Total virus particles (vp) was determined by OD260 and total infectious particles was determined using anti-hexon antibodies against AdHu5 (AdenoX Rapid Titer kit, Clontech) or against PAV3 (VIDO, University of Saskatchewan). Four independent vector preparations were used for each vaccine. Total infectious particles and total viral particles were determined for both adenovirus vaccines, with ratios of 1:285, 1:333, 1:250, and 1:300 for PAV3-HA and 1:150, 1:250, 1:220, and 1:250 for AdHu5-HA.