Figure 6. PMCAb amplifies PrP^CWD from white-tailed deer with allelic variations and from non-CNS tissues.

Brain homogenates (BH; 10% w/v) from CWD-positive deer with allelic variations of Prnp were tested for their ability to seed PMCAb. CWD-positive 10% BH from (A) wt/wt or wt/95H deer and (B) wt/96S or 95H/96S white-tailed deer were serially diluted in normal BH (NBH) from Tg(CerPrP)1536^+/− mice, and 10 µL of the 25-, 625- and 1.6×10⁴-fold dilutions from each were used to seed 90 µL fresh NBH and subjected to one 96-cycle PMCAb round. (C) For testing the homozygous 96S allelic variant, retropharyngeal lymph nodes (LN) or BH from two separate hunter-harvested CWD-positive deer were used to seed NBH for PMCAb. As controls, homogenates of lymph nodes from a hunter-harvested CWD-positive wt/wt deer or orally inoculated wt/wt CWD clinically affected deer was used to seed a 96-cycle round of PMCAb. (D) Immunoblotting three five-fold dilutions of 10% BH from the end-stage wt/wt deer used for these studies after proteinase K digestion demonstrates that the PrP^CWD concentrations used in the PMCAb reactions (Fig. 6A) were not detectable without amplification.