Fig. 1
Biochemical and histopathological strain phenotype of MM2-sCJD prions in human PrP mice. (a) Electrophoretic pattern of cortical MM2-sCJD prions in human brain and in human PrP mouse (tg650) brains and spleens. Tissue homogenates were subjected to western blot analyses after limited proteinase K digestion. Blots were probed with Sha31 antibody. Other human prion sources (MM1-sCJD, E200K familial CJD and variant CJD) are shown as controls. The equivalent of 0.5 and 2 mg of brain and spleen tissue was loaded on the SDS-PAGE gel. Red and blue arrows denote the unglysosylated bands of PrPres migrating at 21 kDa (T1) and 19 kDa (T2), respectively. Molecular masses (MM) of protein standards are indicated in kilodaltons. (b) Ratio of diglycosylated and monoglycosylated PrPres species in the brains of tg650 mice following serial transmission (4 passages) of MM2-sCJD (circles) and MM1-sCJD (squares) prions (data plotted as means ± SEM, n = 6 mice analyzed at each passage). (c) Western blot analysis of PrPres in the brain of tg650 mice infected with MM2-sCJD prions, after blotting with Sha31 antibody (top) or 12B2 antibody specific for Type 1 PrPres (bottom). The banding patterns observed on transmission of other CJD subtypes and atypical L-BSE (which exhibits also a T2 signature, [4]) are shown for comparison. The equivalent of 1 mg (Sha31) and 7 mg (12B12) tissue were run on the SDS-PAGE gels for MM2-sCJD and L-BSE infected brains. The equivalent of 1 mg (Sha31, VV1, MM1, MV1; 12B2, MV2, VV2), 0.5 mg (Sha31 MV2, VV2), 2 mg (12B2, VV1, MM1, MV1) were loaded for the other samples. Note the presence of low-size PrPres fragments in the brain of tg650 mice infected with MV2 and VV2 sCJD sources (black arrow). (d) Western blot analysis of PrPres in the brain of tg650 mice infected with MM2-sCJD prions, after deglycosylation by PNGase F. Blots were probed with Sha31. Other CJD subtypes are shown for comparison. The equivalent of 0.2 mg of brain tissue (MM1, MM2), 0.1 mg (MV1, VV1) and 0.05 mg (MV2, VV2) were run on the SDS-PAGE gels. MM: molecular mass standards. (e) Regional distribution of PrPres in the brain of tg650 mice infected with MM2-sCJD prions, by representative histoblots in 4 different antero-posterior sections. See [7] for comparison with transmission of MM1-sCJD prions. Histoblots were probed with 12F10 anti-PrP antibody