(A) Immunological cross-reactivity of hamster faeces in the Western blot (labeling with mAb 3F4). Lanes 1 and 4: PK digested 263K scrapie hamster brain homogenate containing 5 × 10⁻⁷ g of brain tissue. Lanes 2 and 3: Crude homogenates from 2 mg of normal hamster faeces. Lanes 5 and 6: Purified extracts from 100 mg of normal hamster faeces. (B) Semi-quantitative Western blot assessment of the extraction recovery of PrP27-30, the PK-resistant core of PrP^TSE, from hamster faeces spiked with 263K scrapie brain homogenate. Lanes 1–5, 9–13 and 17–21: Serially diluted 263K scrapie hamster brain homogenate containing 1 × 10⁻⁶ g (lanes 1 and 9), 7 × 10⁻⁷ g (lanes 2 and 10), 5 × 10⁻⁷ g (lanes 3 and 11), 2.5 × 10⁻⁷ g (lanes 4, 12 and 17), 1 × 10⁻⁷ g (lanes 5, 13 and 18), 5 × 10⁻⁸ g (lane 19), 2 × 10⁻⁸ g (lane 20) and 1 × 10⁻⁸ g (lane 21) tissue. Lanes 6-8: Extracts from 200 mg normal hamster faeces spiked with 5 × 10⁻⁶ g of homogenized 263K scrapie brain tissue before purification. Lanes 14-16: Extracts from 200 mg normal hamster faeces spiked with 3 × 10⁻⁶ g of homogenized 263K scrapie brain tissue before purification. Lanes 22–23: Extracts from 200 mg normal hamster faeces spiked with 5 × 10⁻⁷ g of homogenized 263K scrapie brain tissue before purification. (C) Western blot detection of PrP27-30, the PK-resistant core of PrP^TSE, in faeces of hamsters fed with 263K scrapie brain homogenate. Lanes 1–5: Serially diluted 263K scrapie hamster brain homogenate containing 5 × 10⁻⁶ g (lane 1), 1 × 10⁻⁶ g (lane 2), 2.5 × 10⁻⁷ g (lane 3), 1 × 10⁻⁷ g (lane 4) and 2.5 × 10⁻⁸ g (lane 5) tissue. Lanes 6-21: Extracts from 200 mg faeces sampled at 0–24 hpi (lanes 6-9), 24–48 hpi (lanes 10-13), 48–72 hpi (lanes 14–17) and 72–96 hpi (lanes 18–21) from 4 different animals. (D) Western blot detection of PrP27-30, the PK-resistant core of PrP^TSE, in faeces of hamsters fed with 263K scrapie brain homogenate – Adjustment of dilutions for semi-quantitative assessment of PrP^TSE excretion. Lanes 1–5: Serially diluted 263K scrapie hamster brain homogenate containing 5 × 10⁻⁷ g (lane 1), 2.5 × 10⁻⁷g (lane 2), 1 × 10⁻⁷ g (lane 3), 5 × 10⁻⁸g (lane 4) and 2 × 10⁻⁸ g (lane 5) tissue. Lanes 6–9: 1:2 (lanes 6 and 8) and 1:5 (lanes 7 and 9) diluted extracts from 200 mg faeces sampled at 24–48 hpi (dilutions were prepared from the extracts represented in lanes 10 and 12 of Fig. 2C). (E) Western blot detection of PrP27-30, the PK-resistant core of PrP^TSE, after serial protein misfolding cyclic amplification (sPMCA) of PrP^TSE in faeces extracts from clinically diseased scrapie hamsters fed with 263K scrapie agent. Lane 1: 263K scrapie hamster brain homogenate containing 5 × 10⁻⁷ g tissue. Lanes 2–11: sample signals of extracts from 200 mg faeces of two donor animals after 200 (lanes 2 and 3), 250 (lanes 4 and 5), 300 (lanes 6 and 7), 350 (lanes 8 and 9) and 400 (lanes 10 and 11) cycles of sPMCA.