Tissues from all three patients with mutations in the human PrP gene (PRNP) were obtained from Dr. Annemieke Rozemuller at the Dutch Surveillance Center for Prion Diseases, University Medical Center Utrecht (UMCU), Utrecht, The Netherlands and the VU University Medical Center in Amsterdam, The Netherlands. Frozen brain tissue from the 55-year old Y226X mutant patient (UMCU #S08–005)  was not available. Therefore, brain tissue from the cingulate gyrus was provided as formalin-fixed (three days), paraffin-embedded tissue sectioned and dried on glass slides. To create the brain homogenate used for inoculation into mice, tissue from 10 slides (approximately 1 cm2 each) was de-paraffinized and rehydrated using standard protocols, and then scraped from the slides using a razor blade. The resulting material was minced into small pieces (less than 0.5 mm2) with a scalpel and added to a total volume of 500 μl phosphate buffered saline (PBS). This suspension was vortexed and sonicated for several rounds of 30 s each until no large pieces could be observed.