Immunohistological (A-L) and immunoblotting (M-N) results. A C: frontal cortex; D F: temporal cortex; G L: cerebellum. Spongiform change in the frontal (A) and temporal (D) cortex and molecular layer of the cerebellum (G, H) is accompanied by moderate neuronal loss in cortex (A, D) and torpedoes in the granular layer of the cerebellum (I). PrP-immunoreactive (PrP-ir) deposits are seen in the cerebral cortex and cerebellum (B, E, J). PrP-ir is largely reduced in the cerebral cortex after proteinase K (PK) treatment, except for small PrP-ir dots following a dot-like or target-like pattern (C, F). By contrast, PrP-ir in the molecular layer of the cerebellum, in the form of elongated plaque-like deposits, is preserved after PK treatment (K, L); PrP plaques in the granular layer are absent. Paraffin sections: A, D, G, H: hematoxylin and eosin staining; I: phosphorylated neurofilament immunohistochemistry; B, C, E, F, J L: PrP immunostaining (3F4 antibody) without (B, E, J) and with (C, F, K, L) PK treatment. A, D, G, J, K, L: × 200 (bar in L, 100μm); B, C, E, F, H, I: × 400 (bar in I, 50μm). PK was used according to the indications of the supplier: 1 drop of PK concentrate (DAKO, S2019) in 1.6mL of DAKO ChemMate TM PK diluent (S2032) for 15 minutes. M: Routine immunoblotting conditions (10% brain homogenate and final PK concentration of 440μg/mL) as described elsewhere  and five minutes of film exposure time. PK pretreated brain regions corresponded to occipital cortex (lane 1), putamen/globus pallidus (lane 2), cerebellum (lane 3), parietal cortex (lane 4), thalamus (lane 5), frontal cortex (lane 6), temporal cortex (lane 7), sporadic Creutzfeldt–Jakob disease (sCJD) VV2 reference case occipital cortex (lane 8). N: Immunoblotting of PK pretreated samples with less stringent conditions (TeSeE® Western Blot Kit, Bio-Rad) and detection with 3F4 antibody (Dako, dilution 1:3000) as previously described  at ten minutes film exposure time. Brain regions corresponded to occipital cortex (lane 1), cerebellum (lane 2), parietal cortex (lane 3), thalamus (lane 4), frontal cortex (lane 5), temporal cortex (lane 6), variably protease-sensitive prionopathy 129MV parietal cortex (lane 7)  and sCJD VV2 reference case frontal cortex (lane 8). Molecular weight standards are indicated in kDa: (M) SDS-PAGE Standards, broad range, Bio-Rad and (N) Precision Plus Protein Unstained Standards, Bio-Rad.