Figure 6

Immunoblot screening for PrPres in CWD-inoculated tg66 and tgRM mice. Brains from CWD-inoculated transgenic mice were treated with proteinase K (PK) or untreated and processed for immunoblot as described in the methods. A, B Results without PTA precipitation, C, D used PTA precipitation to concentrate potential low levels of PrPres. CWD inoculum and mouse numbers are shown across the top of each immunoblot. PK and PTA treatment status are shown across the bottom. A (tg66 mice): lane 1, tg66 mouse infected with vCJD; lanes 2, 3 uninoculated tg66 mouse; lanes 4–11 show several tg66 CWD-inoculated mice, including two RT-QuIC suspect mice (bolded). B (tgRM mice): lane 1, tg66 mouse infected with vCJD; lanes 2, 3 uninoculated tgRM mouse; lanes 4–11 show several tgRM CWD-inoculated mice. For both A and B, lane 1 was loaded with 0.36 mg tissue equivalents (te), lanes 2 and 4–11 were loaded with 0.72 mg te each and lane 3 was loaded with 0.04 mg te. C, D (PTA precipitation of tg66 mice): lane 1, tg66 mouse infected with vCJD; lanes 3, 4 uninoculated, age-matched tg66 mice; lanes 5–10 in C and 5–9 in D show several tg66 CWD-inoculated mice, including two RT-QuIC suspect mice (bolded). Lane 10 in D shows PTA precipitated PrPsen from an uninfected tg66 mouse. For C and D, lane 1 was loaded with 0.24 mg te, lanes 2–10 in C and lanes 2–9 in D were loaded with 22.7 mg te and lane 10 in D was loaded with 1.35 mg te. The immunoblots were probed with anti-PrP antibody 3F4 and developed using a femto detection system.