Immunoblot analysis of abnormal PrP propagated in the brains of 117VV Tg30 transgenic mice challenged with vCJD prions.

<p>All mice were inoculated with the same vCJD prion isolate. Immunoblots were analysed by enhanced chemiluminescence with monoclonal anti-PrP antibody ICSM 35. The provenance of the brain samples is designated above the lanes. (A) Lanes 3 and 4 show the predicted type 5 PrP^Sc bands seen when vCJD is propagated in transgenic mice expressing HuPrP with the codon 129VV genotype, compared with the detection of type 4 PrP^Sc in the brain of vCJD-inoculated 129MM Tg45 mouse (lanes 1 and 2). Type 5 shares the glycoform ratio of type 4 but differs in migrating more slowly on western blots because all 3 glycoform fragments of type 5 have higher apparent molecular masses than those of type 4. The lower signal intensity in lane 3 (100 µg/ml PK) compared to lane 4 (50 µg/ml PK) reflects PK-sensitivity of the vCJD-seeded 117V PrP^Sc. The 8 kDa PrP fragments can be seen associated with only PK-digested prion-infected 117V PrP-expressing mouse brain samples. These truncated 8 kDa fragments are absent from either vCJD infected Tg45 mice (lanes 1 and 2) or vCJD-infected 117VV Tg30 brain samples not digested with PK (lane 5).These data confirm that the 8 kDa human PrP fragment is a disease-associated PrP degradation product. (B) Variable PK resistance in brain of 117VV Tg30 mouse inoculated with vCJD. The 8 kDa PrP fragment is only seen in PK digested lanes 1–4, but is absent in the same sample undigested with PK (lane 5).