NR AWPM
AU Ushiki,Y.K.; Endo,R.; Shimizu,Y.; Iwamaru,Y.; Yamamoto,T.; Hattori,S.; Itohara,S.; Shinagawa,M.; Irie,S.; Yokoyama,T.
TI Monitoring of conformational transition of PrPsc during interspecies transmission by using novel PrPsc-specific antibodies
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-27
PT Konferenz-Poster
AB A panel of PrPsc conformation-specific antibodies can be valuable tools in the structural analysis of PrPsc. In order to develop novel PrPsc-specific monoclonal antibodies (mAbs), we immunized PrPdeficient mice with PrPsc that was purified from scrapie-infected mouse brains. The brain homogenate of a scrapie-infected mouse was coated onto an ELISA plate and incubated with (denatured) or without (native) guanidine prior to hybridoma screening. In this screening, we selected candidate mAbs that could act as PrPsc-specific mAbs; these mAbs exhibited stronger immunoreactivity toward the native scrapie brain homogenate than toward the denatured homogenate. Further, each mAb was characterized by immunoprecipitation assays. Of all the tested mAbs, 3B7 exhibited immunoreactivity against PrP in the brain homogenate of the scrapie-infected mouse, but not against that in the brain homogenate of a normal mouse. This result may confirm its specific immunoreactivity against PrPsc. This mAb also reacted with PrPsc in the brain of scrapie-infected hamster, but not with PrPsc in the brain of scrapie-infected sheep and cattle suffering from bovine spongiform encephalopathy. This result showed that there is conformational difference in the PrPsc of rodents (mice and hamsters) and ruminants (sheep and cattle). Using the mAb 3B7, we examined the conformational transition of PrPsc during interspecies transmission. The immunoreactivity of the mAb 3B7 against PrPsc in the brains of scrapie-infected sheep and sheep scrapie-passaged mice (primary to 5th passage) was examined by an immunoprecipitation assay. No signal was detected from the PrPsc in primary and secondary passaged mice brains; this was also observed in the case of scrapie-infected sheep brains. However, the mAb 3B7 could react with PrPsc in mice brains after the 3rd passage. Based on the length of the incubation periods, prion adaptation requires at least 3 passage histories for interspecies transmission. This conformational change might be correlated with a decreased incubation period. Our data provides direct evidence for the conformational transition of PrPsc during prion adaptation.
AD Y.K. Ushiki, T. Yamamoto, S. Hattori, S. Irie: Nippi Research Institute of Biomatrix, Tokyo, Japan; R. Endo, Y. Shimizu; Y. Iwamaru, M. Shinagawa: National Institute of Animal Health, Tsukuba, Japan; S. Itohara: Brain Science Institute, RIKEN, Wako, Japan
SP englisch
PO Italien