Three mite species (Demodex ursi, Ursicoptes americanus, and Sarcoptes scabiei) have been associated with mange in black bears (Ursus americanus). Since the early 1990s, the number and geographic distribution of mange cases in black bears in Pennsylvania, US has increased; however, the causative mites have yet to be completely defined. We evaluated several diagnostic approaches for detection and identification of mites in 72 black bears with severe lesions consistent with mange. Sarcoptes scabiei was morphologically identified in skin scrapes from 66 of the bears; no mites were identified in the remaining six. Histopathologic lesions consistent with sarcoptic mange were observed in 39 of 40 bear skin samples examined, and intralesional mites were observed in samples from 38 of these bears. Samples were collected from a subset of the 72 bears for PCR testing targeting both the internal transcribed spacer (ITS)-2 region and cytochrome c oxidase I (cox1) gene including 69 skin scrapes (ITS-2 only), 56 skin biopsies (ITS-2 and cox1), and 36 fecal samples (ITS-2 and cox1). Skin scrapes were a more sensitive sample for PCR detection than either skin biopsies or fecal samples, and the ITS-2 primers proved more sensitive than cox1. Using a commercial indirect enzyme-linked immunosorbent assay, antibodies to S. scabiei were detected in 45/49 (92%) black bears with confirmed mange and 0/62 (0%) cubs with no gross lesions suggestive of mange and which were born to seronegative sows. Sarcoptes scabiei was the predominant mite associated with mange in black bears in Pennsylvania. Diagnostically, cytologic examination of skin scrapes was the most effective approach for diagnosing active mite infestations in black bears. The evaluated serologic assay accurately detected antibodies to S. scabiei in most bears with confirmed S. scabiei infestations. Additional research is needed to determine the usefulness of this approach for larger scale surveys and for asymptomatic bears.