Feeding schedule for bioassay in Syrian hamsters

*<p>mg relates to soil sample; µl relates to aqueous extract</p

Western blot analysis of hamsters orally challenged with contaminated soil and western blot typing of PrP27-30.

<p>a) Western blot showing PrP^Sc in the brains of hamster orally challenged with contaminated soil samples and in the soil samples used for the bioassay. Lane 1: negative control hamster; lane 2: hamster H19 fed 12 times with aqueous extracts of soil samples from month 26 and 29; lane 3: hamster H4B fed 12 times with soil samples from month 26 and 29; lane 4: scrapie-contaminated soil (18 months); lane 5: scrapie-contaminated soil (21 months); lane 6: negative soil sample. b) Western blot typing of electrophoretic mobilities and glycosylation characteristics of PrP27-30 from different hamster-adapted TSE reference isolates, and from hamsters perorally challenged with 263K scrapie-contaminated soil. Lane 1: ME7-H scrapie agent; lane 2: hamster-adapted BSE-isolate (BSE-H); lane 3: 263K scrapie agent, lane 4: H10 (hamster 10, fed 12 times with soil samples from months 26 and 29); lane 5: H7 (hamster 7 fed 12 times with soil samples from month 26 and 29).</p

Western Blot Detection of PrP27–30 in Skin Specimens of Hamsters Intracerebrally Challenged with BSE-H Agent

<p>Lanes 1 and 2: skin samples from hindlimb and forelimb of a BSE-infected hamster. Amounts of tissue represented in lanes: 1, 43 mg; 2, 58 mg.</p

Time-Course of PrP^Sc Deposition in Skin Tissue

<div><p>(A–E) Western blot detection of PrP27–30, the protease-resistant core of PrP^Sc, extracted from different skin samples of hamsters orally challenged with 263K scrapie and sacrificed at the following time-points after infection: (A) 70 dpi, (B) 100 dpi, (C) 130 dpi, (D) at the onset of clinical signs for scrapie (138–146 dpi), and (E) at the terminal stage of disease (157–171 dpi). Lanes with test samples: S1, skin sample from hindlimb; S2, skin sample from forelimb; S3, skin sample from back; S4, skin sample from abdomen; S5, skin sample from head. Lanes with control samples: 1, proteinase K–digested brain homogenate from terminally ill 263K scrapie hamsters containing 1 × 10⁻⁷ g brain tissue. Representative results are shown for each stage of incubation. Substantial individual variation was observed at 130 dpi, with two of five and three of five animals displaying findings as in (C) in the Western blot on the left-hand side or the Western blot on the right-hand side, respectively.</p><p>(F) Lanes S1d–S5d: Same samples as in S1–S5 of (E) but deglycosylated with PNGaseF.</p><p>(A–F) Amounts of tissue represented in lanes: (A) S1, 43 mg; S2, 52 mg; S3, 68 mg; S4, 58 mg; S5, 73 mg; (B) S1, 78 mg; S2, 44 mg; S3, 63 mg; S4, 67 mg; S5, 50 mg; ([C], Western blot on the left side) S1, 42 mg; S2, 76 mg; S3, 61 mg; S4, 58 mg; S5, 73 mg; ([C], Western blot on the right side) S1, 51 mg; S2, 63 mg; S3, 70 mg; S4, 87 mg; S5, 54 mg; (D) S1, 63 mg; S2, 68 mg; S3, 90 mg; S4, 50 mg; S5, 68 mg; (E) S1, 55 mg; S2, 73 mg; S3, 80 mg; S4, 88 mg; S5, 70 mg; (F) S1d, 12 mg; S2d, 14 mg; S3d, 19 mg; S4d, 12 mg; S5d, 20 mg.</p><p>(G) Time-scale displaying the mean incubation period and the pre-clinical and clinical phases of incubation of hamsters orally infected with 263K scrapie. Small vertical arrows indicate time-points at which animals were tested for PrP^Sc in skin samples.</p></div

PrP^Sc in Skin Samples from Different Body Areas of Sheep Naturally Infected with Scrapie

<p>Western blot detection of PrP27–30, the protease-resistant core of PrP^Sc, using the anti-PrP antibodies ICSM-18 (lanes 1–8, 10–12) and P4 (lane 9). Samples are from sheep Sc3 (lanes 1–5), sheep Sc5 (lanes 6–9), and from an uninfected control sheep. Lanes 1 and 2: different amounts of brain homogenate (0.5 mg/lane and 0.1 mg/lane of midbrain tissue, respectively) from sheep Sc3. Lane 3: tonsil sample. Lane 4: tonsil sample after deglycosylation with PNGaseF. Lane 5: skin sample from the inguinal region. Lane 6: skin sample from the perianal region (scratching area). Lane 7: skin sample from the same region as in lane 6 after deglycosylation with PNGaseF. Lane 8: skin sample from the snout. Lane 9: skin sample from the same region as in lane 8, but detected with the antibody P4. Lanes 10–12: negative controls of skin samples from different body regions of an uninfected sheep. Amounts of tissue represented in lanes: 1, 0.5 mg; 2, 0.1 mg; 3, 7 mg; 4, 2 mg; 5, 88 mg; 6, 89 mg; 7, 23 mg; 8, 90 mg; 9, 92 mg; 10, 80 mg; 11, 87 mg; 12, 94 mg.</p