No evidence of proteinase K (PK)-resistant prion protein in bighorn sheep brain tissues. (A) Individual bighorn sheep (BHS) were given unique letters to identify them (A-L). The obex region of the medulla from each animal was collected, homogenized to 10% w/v, a 25 μL aliquot of each homogenate incubated in the presence of 50 μg·ml-1 of proteinase K (+) or in the absence of PK (−) and then immunoblotted. Samples from the other BHS in our tissue library similarly did not have PK-resistant prion protein. (B) A dilution series of classical scrapie brain homogenate was prepared in BHS brain homogenate, incubated in the presence of 50 μg·ml-1 of proteinase K and immunoblotted. Scrapie dilution factors were 3, 10, 30, 100, 300 and 1000 (lanes 1–6). Lanes 7 and 8 represent non-proteinase K-treated and proteinase K-treated BHS brain homogenate in the absence of scrapie brain homogenate, respectively. Immunoblots used monoclonal antibody BAR 224.