Fig. 6
RCA can detect rSDS-PrPSc oligomers at early stages of the disease. a Fifty microliters of 10 % hamster brain homogenates (NBH or infected with the 263 K prion strain) were diluted in 300 μL of PBS/2 % Sarkosyl, incubated with 1.5 mM MR100 at room temperature for 1 h and then centrifuged at 8000 g for 5 min. Supernatants (S) were collected and 30 μL of each supernatant were mixed with an equivalent volume of 2X loading buffer. Pellets (P) were resuspended in 30 μL PBS/2 % Sarcosyl, and mixed with 30 μL of 2X loading buffer. Thirty microliters of each sample were loaded on 12 % Bis-tris gels (Criterion, Biorad) and western blotting was carried out with the SAF mix according to standard procedures [16]. Molecular masses (20–75 kDa) are indicated on the left side of the panels. b Hamster brain tissues were collected at various days post-infection (d.p.i.), as indicated, and freshly homogenized tissues were processed according to the RCA protocol using MR100 and analyzed by immunoblotting as described above. c To compare the RCA and the PK test, the same hamster brain homogenates (at 109, 130 and 148 d.p.i.) were incubated with MR100 at room temperature for 1 h, then digested with 20 μg/mL of proteinase K and processed as described in the legend to Fig. 4a. The asterisk in b and c indicates the position of oligomer traces