NR AWHU
AU Karino,A.; Furuoka,H.; Kimura,K.; Shinagawa,M.; Horiuchi,M.
TI Generation of MAB that distinguishes PrPsc from PrPc and neutralizes prion infectivity
QU International Conference - Prion 2006: Strategies, advances and trends towards protection of society - 3.10.-6.10.2006, Torino, Italy, Lingotto Conference Centre - Poster sessions S-11
PT Konferenz-Poster
AB PrPsc-specific molecular probes, such as monoclonal antibody (mAb), are indispensable tool for elucidating the entity of prion. To stablish PrPsc-specific mAbs, we immunized PrP-/- mice with PrPsc purified from prion-infected mice, and screened mAbs with purified PrPsc. Finally we obtained mAb 6H10 that appeared to be a candidate for the PrPsc-specific mAb. The 6H10 reacted with PrPsc treated with proteinase K, whereas the reactivity of pan-PrP mAbs to PrPsc was disappeared under the same condition. The 6H10 did not react with PrPsc pretreated with more than 3M GdnHCl, while the reactivity of pan-PrP mAbs increased with the increase of GdnHCl concentration used for pretreatment of PrPsc. In histoblot analysis, the 6H10 showed positive reaction to non-denatured histoblot but the reactivity decreased after the denaturation of histoblot by autoclaving. In contrast, the reactivity of pan-PrP mAb was enhanced after autoclaving. These results suggested that 6H10 recognizes a conformational epitope on PrPsc. Peptide phage display analysis suggested that the extreme C-terminus of PrP may be involved in constituting the epitope for 6H10. The 6H10 immunoprecipitated PrPsc from prion-infected brains of mouse, sheep, and cattle. Furthermore, pretreatment of purified PrPsc with 6H10 reduced the infectious titer by 1Log10 in mouse bioassay. However, 6H10 showed weak reaction to brains of uninfected Prnp+/+ and Prnp-/- mice in histoblot analysis, suggesting that 6H10 has cross-reactivity to other host factor(s) than PrP. Nevertheless, the mAb 6H10 will be a useful tool for analyzing the relationship between biochemical and biological properties of prion.
AD A. Karino: Department of Veterinary Public Health, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Japan; H. Furuoka: Department of Veterinary Pathology, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Japan; K. Kimura, M. Shinagawa: Prion Disease Research Center, National Institute of Animal Health, Tsukuba, Japan; M. Horiuchi: Laboratory of Prion Diseases, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo, Japan. E-mail: horiuchi@vetmed.hokudai.ac.jp
SP englisch
PO Italien