Acta Neuropathol (2009) 118:659–671 Table 6 Nomenclature and classiﬁcation of sCJD subtypes Nomenclaturea %b Pure subtypes MM/MV 1; VV2; MV 2K; MM/MV 2C; MM 2T; VV1; Mixed subtypes MM/MV 1?2C MM/MV 2C?1 VV 2?1 MV 2K?1 MV 2 K?C MM 2 T?C 65 26 2 3 1 3 \1 669 Distinctive histopathologic features Previously established [30] (for a summary see Table 8 in [30]) As in MM/MV 1 but with clusters of large vacuoles associated to perivacuolar and coarse PrP deposition mainly in cerebral cortex or thalamus As in MM/MV 2C but with synaptic-type PrP staining in the molecular layer of the cerebellum Virtually indistinguishable from VV2 Virtually indistinguishable from MV 2K As in MV 2K but with clusters of large vacuoles associated to perivacuolar and coarse PrP deposition mainly in cerebral cortex As in MV 2T but with clusters of large vacuoles associated to perivacuolar and coarse PrP deposition mainly in cerebral cortex a It is largely based on codon 129 PRNP genotype, which can be either methionine (M) or valine (V) and the PrPSc type (1 or 2 according to Parchi et al. [28, 29]). Since both MM 2 and MV 2 groups are associated to 2 distinct phenotypes, these are further deﬁned with a third parameter (capital letter) referring to distinctive histopathological features: K kuru type amyloid plaques, C predominant cortical pathology with conﬂuent vacuoles and perivacuolar PrP staining, T prominent thalamic pathology with atrophy b Percentage of total consecutive sCJD cases (n = 200) investigated distinct subtype in terms of biological relevance, it is important that they are properly identiﬁed and are added as new subtypes in the current sCJD classiﬁcation (Table 6). Despite the emphasis on molecular features of current sCJD classiﬁcation, it has become increasingly clear that PrPSc typing alone, when limited to a single or even a few brain samples, fails to provide an accurate classiﬁcation in a signiﬁcant proportion of cases. This is mainly related to the focal nature of the ‘‘mixed features’’ in many sCJD cases with PrPSc types 1 and 2 concurrence. Indeed, we would have misclassiﬁed the disease subtype in about 27.5% (using 3F4) of cases with MM or MV genotype, if we had analyzed PrPSc only in the frontal cortex, the area more commonly used for typing worldwide. For the same reason, discrepancies may arise when PrPSc-typing and PrP immunohistochemistry are performed from individual samples taken from opposite hemispheres or even adjacent cortical gyri. However, our study shows that the regional deposition of either type 1 or type 2 when concurrent is not random and that a relatively limited number of critical brain structures must be assessed to reach an accurate classiﬁcation. Furthermore, our results further underline the importance of applying both molecular and neuropatho- logical assessment for sCJD subtype classiﬁcation. In this regard, the lack of detection of PrPSc type 2 in a minority of MM subjects, despite the presence of a mixed synaptic and perivacuolar pattern of PrP deposition, indicates that when type 2 is very focal or limited in amount, histopathologic examination is more sensitive in identifying such cases than PrPSc typing, at least when only the 3F4 antibody is used. Given the very strong correlation in MM subjects between PrPSc type 2 detection and the large ‘‘grape-like’’ vacuoles and the perivacuolar pattern of PrP deposition on histopathologic examination, which is in line with results previously obtained in other studies [18, 37], we propose that these cases are classiﬁed as MM 1?2C or MV 1?2C even without the ﬁnal proof of type 2 detection by western blot. Alternatively, PrPSc typing using the antibody 1E4 was in our hand as sensitive as the histopathologic exam- ination in the detection of cases with very focal type 2. In the light of the present results, the most important regions to be assessed pathologically include the cerebral cortex from each of the 4 lobes, the striatum, hippocampus, thalamus and cerebellum. The cerebellum, in particular, is critical for the recognition of the synaptic pattern of PrP deposition as marker of PrPSc type 1 concurrence in the cases with dominant type 2. Taken together, our data indicate that a protocol including the neuropathologic assessment of the eight brain regions mentioned above and PrPSc typing in four critical regions such as the temporal, parietal and occipital neo- cortices, and medial thalamus is strongly recommended for a reliable sCJD group classiﬁcation addressing the issue of mixed phenotypes. Indeed, by applying this protocol 123