Samples of frozen tissues were thawed, weighed and homogenized in phosphate buffered saline, pH 7.4 (PBS) to give 10% (brain) or 20% (lymphoid tissues) w/v homogenates. Western blotting for PrPSc detection was performed following proteinase K digestion and sodium phosphotungstic acid (NaPTA) precipitation of tissue homogenates, as previously described [27], using the primary anti-PrP monoclonal antibodies ROS-BC6 (raised against recombinant sheep PrP residues 94–233; epitope amino acids 144FGNDYEDRYYR154) and ROS-IH9, both at 0.5 μg/ml [25].

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