In a retrospective study, immunohistochemistry was applied to 37 archival cases of rabies using a commercial polyclonal antibody. Thirteen different species were studied including: 3 dogs, 4 cats, 1 pig, 5 cattle, 3 horses, 1 llama, 7 skunks (Mephitis mephitis), 7 raccoons (Procyon lotor), 1 bat (Myotis sp.), 1 white-tailed deer (Odocoileus virginianus), 1 bobcat (Lynx rufus), 2 gray foxes (Urocyon cinereoargenteus), and 1 red fox (Vulpes vulpes). All cases had previously been diagnosed as rabies using histopathology and/or fluorescent antibody testing. The immunohistochemistry technique successfully detected the presence of rabies virus antigen in every case, and highlighted characteristic distributional differences throughout the brain for most species. The results showed the hippocampus as the best site for rabies detection in both dogs and cats. For cattle, the virus particles were most prominent in the brainstem, followed by the cerebellum. In horses, the cervical spinal cord and adjacent brainstem proved to be optimal sites for detecting rabies. In raccoons and skunks, positive labeling was widely dispersed and selection might be less important for these wildlife reservoir species. Immunohistochemistry should prove useful in enhancing the accuracy of rabies diagnosis through informed selection of brain segments when composite sampling is not feasible. This technique, which uses formalin-fixed tissue, has several advantages over fluorescent antibody testing which is performed on fresh tissue; specifically, it avoids any biosafety hazards in transport or in the laboratory, and also allows for further histological examination. Such widespread success of the commercial antibody shows great promise for becoming a universal test for identifying different rabies virus variants in any mammalian species. Also, by making immunohistochemistry available for diagnostic laboratories in the developing world, more rapid and reliable virus detection could help to reduce the high number of human rabies deaths prevalent in these areas.