Antemortem and postmortem evaluations revealed no evidence of prion infection in three captive mountain lions (Puma concolor) consuming chronic wasting disease–infected carcasses for ≥14.5 yr. (A) Neurologic assessments by operant conditioning (Supplementary Material Table S1) revealed no clinical signs suggestive of progressive encephalopathy or other neurologic disease. (See Supplementary Video S2 of representative assessments.) (B) Intense immunohistochemistry (IHC) stain in cecal contents but not in gut-associated lymphoid tissue. Bar=200 µm; IHC with monoclonal antibody (mAb) F99/97.6.1 (slide 12, report WHL 16 949 in Supplementary Material). (C–E) A few areas of mild vacuolation (C) were observed (pictured: external cuneate nucleus of the brainstem; H&E stain), but no immunostained PrP^d accumulations after IHC staining with either mAb F99/97.6.1 (D) or L42 (E). Bar=50 µm in panels C–E, all from the same section of external cuneate nucleus (slide 7, WHL 16 949).