This study involved the isolation and evaluation of immunizing subunits of bovine adenovirus (BAV) types 2, 3 and 7. These serotypes were chosen as possible candidates to construct a BAV vaccine because they have been implicated in disease in the United States;Before vaccines involving synthetic or recombinant DNA technology can be developed, the immunogen needs to be identified. Monospecific sera against protein subunits of BAV types 2, 3 and 7 were produced in rabbits using agarose immunoprecipitates prepared by preparative line or crossed immunoelectrophoresis with hyperimmune type-specific sera. The specificities of the sera produced were determined by immunoprecipitation/sodium dodecylsulfate polyacrylamide gel electrophoresis, Western blot and immunodiffusion. The viral neutralization potentials of these sera were analyzed using homologous and heterologous assays. In addition, hexon subunits of BAV-3, produced by deoxycholate and heat treatment of virions followed by separation on a DEAE BioGel A column, were used to product hyperimmune sera. Results indicated that the anti-hexon subunits sera seem to be superior to the monospecific sera produced from agarose immunoprecipitates. These hexon subunits were capable of inducing both subgroup- and type-specific viral neutralizing antibodies, thus making them ideal candidates for multivalent subunit vaccines;Before recombinant DNA technology can be used to manufacture immunizing subunits, the relative locations of the genes coding for the major capside proteins need to be identified on the genome. Cross-hybridization studies were performed on human adenovirus type 2 (HAV-2) and BAV-7 DNAs, using cloned restriction fragments as probes. Under normal conditions (0.4 NaCl, 65(DEGREES)C), no hybridization was detected. However, at reduced stringency (1M NaCl, 57(DEGREES)C), two distinct regions of homology were observed. One region centered around 10.3 to 15.5 map units (m.u.) on the HAV-2 genome, which codes for the IVa2f protein. The other region lay between 41 to 70 m.u. on HAV-2, with the most conserved region (50.1 - 58.5 m.u.) coding for the hexon protein. These results allow the alignment of the genomes of the two viruses.