Figure 6

Exogenous cellular co-factors do not modify the efficiency of M1000 infection in vitro or in vivo. (A) Representative cell blot showing PrP^res levels produced by recipient MoRK13 cells exposed to M1000 brain homogenate diluted to three different final concentrations (as indicated) with either 1:4 lysis buffer (LB):medium or 1:4 MoRK13 fraction:medium mix. (B) Quantification of PrP^res produced by MoRK13 cells exposed to M1000 brain homogenate diluted in subcellular fraction relative to the equivalent % M1000 brain homogenate diluted in lysis buffer; data is from M1000 spiked into three independent fractionations of MoRK13. For quantification purposes, the three different % M1000 spikes into the same MoRK13 fractionated lysate were considered a triplicate of the same experiment. Fractions enriched in lipid raft (LR), endoplasmic reticulum (ER), mitochondrial (MT) and early endosome (EE) marker proteins are highlighted. Analysis of relative levels of PrP^res produced after infection was by one-way ANOVA with Tukey's multiple comparison test; no significant differences found comparing subcellular fractions. (C) Survival of Tga20 mice inoculated with M1000 brain homogenate diluted to 0.01% with either PBS, selected fractions from an empty Nycodenz gradient, or selected subcellular fractions from MoRK13 cells. Each data point represents the number of days post-inoculation (PI) which mice were sacrificed because of terminal prion disease. Statistical analysis was by one way ANOVA, with no significant differences in survival seen between any inocula.