Epitope analysis.

<p>(A) The epitopes of 6A12 and 8D5 in the PrP amino acid sequence. The epitope positions of mAbs 6A12 and 8D5 are represented by boxes under the amino acid sequence of mouse PrP (residues 1–254) (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0058013#pone.0058013.s002" target="_blank">Supplemental Fig. 2</a>). The dashed line indicates the octapeptide repeat region. Sequence numbers are shown on the right. (B) Peptide competition assay. P31–39 and P41–47 are synthetic peptides. MAbs were pre-incubated with (P31–39 or P41–47) and without (−) synthetic peptide prior to use in immunoprecipitation. The brain homogenate [0.05% (w/v)] from BSE-affected mice was incubated with the antibody-peptide complex and immunoprecipitated with protein G-coupled magnetic beads. The immunoprecipitated PrP was western blotted and detected with HPR-conjugated mAb T2. Total PrP in the brain homogenate was detected by routine western blotting (WB). Molecular weight markers are shown on the left (kDa). P31–39 blocked the reactivity of mAb 8D5 to PrP^Sc, but not the reactivity of mAb 6A12 to PrP^Sc. Conversely, P41–47 blocked the reactivity of mAb 6A12 to PrP^Sc, while P31–39 did not.