Western Blot Analysis of Brain PrP Levels

Brains samples were homogenised in NP-40 lysis buffer (1% NP-40, 0.5% sodium deoxycholate, 150 mM NaCl, 50 mM Tris-HCL [pH 7.5]) at 10% weight/volume, and subsequently treated at 37 °C for 1 h with proteinase K (20 µg/ml). Samples were then separated by electrophoresis and electroblotted onto polyvinylidene difluoride membranes as described previously^{42 (link)}. PrP was detected using anti-mouse PrP mAb (clone 7A12; Yin et al., 2007), followed by HRP-conjugated anti-mouse antibody (Jackson Immunoresearch, Ely, UK) and visualised with BM Chemiluminescent substrate kit (Roche, Burgess Hill, UK). An uncropped image of the immunoblot is provided in Supplementary Fig. S1.