Figure 2.

Coronally sliced whole brain prion organotypic slice culture assay (POSCA) can propagate three rodent-adapted scrapie strains. (A) Immunoblots of proteinase K (PK)-digested PrP from slice homogenates of anterior (1) through posterior (10) cultured brain regions. Slices were infected with 22L, RML, or ME7 strain of rodent-adapted scrapie, then harvested at day 56 post-infection. All strains can be propagated in this system. Whether the different levels of PK-resistant PrP are strain-specific is the question of ongoing experiments. POSCA was prepared from C57Bl6 mice for all strains, with the same baseline PrP^C levels. An amount of 50 µg total protein before PK digestion was loaded. (B) Anatomy of a slice from region 6 and heat maps for total PrP (PrP^C plus PrP^Sc) of tga20* POSCA uninfected or infected with 22L and RML, showing different distributions of total PrP. Heat map values indicate relative amounts (percentiles of total PrP) within a slice, not absolute values of PrP, so it is the pattern of total PrP distribution, not the intensity of signal, that can be compared between slices. Unlike immunoblots which only provide total levels of a protein from a homogenate, slice culture allows analysis of regional variation within a single slice. * tga20 mice express wildtype mouse PrP at 6× fold higher levels and were the original mouse line used for the first POSCA experiments. PrP antibodies: Sha31 (A), SAF83 (B).