title: Efficient in vitro amplification of a mouse-adapted scrapie prion protein.
abstract: Protein misfolding cyclic amplification (PMCA) is a highly sensitive technique used to detect minute amounts of scrapie prion protein (PrP(Sc)), a major protein component of the infectious agents associated with prion diseases. Although exponential in vitro amplification of hamster scrapie PrP(Sc) has been established, the PMCA used was unsuccessful in achieving good amplification of PrP(Sc) from other animals. Here, we have investigated the cause of the insufficient PrP(Sc) amplification in mice and have developed an improved method suitable for amplification of the PrP(Sc) of the mouse-adapted scrapie prion strain Chandler. Mouse PrP(C), the cellular form of the prion protein, tends to become resistant to proteases during incubation independent of sonication. By adding digitonin to the reaction buffer as a lipid detergent, accumulation of the protease-resistant PrP(C) was inhibited; hence, mouse PrP(Sc) could be amplified to infinite levels. The present study is the first report describing effective amplification of PrP(Sc) of the mouse-adapted scrapie prion and this improved PMCA technique will contribute to prion research that uses mice as experimental animals.
authors.affiliation: Prion Disease Research Center, National Institute of Animal Health, 3-1-5 Kannondai, Tsukuba, Ibaraki 305-0856, Japan. ymura@affrc.go.jp
authors.forename: Yuichi
authors.initials: Y
authors.lastname: Murayama
authors.forename: Miyako
authors.initials: M
authors.lastname: Yoshioka
authors.forename: Takashi
authors.initials: T
authors.lastname: Yokoyama
authors.forename: Yoshifumi
authors.initials: Y
authors.lastname: Iwamaru
authors.forename: Morikazu
authors.initials: M
authors.lastname: Imamura
authors.forename: Kentaro
authors.initials: K
authors.lastname: Masujin
authors.forename: Sachiko
authors.initials: S
authors.lastname: Yoshiba
authors.forename: Shirou
authors.initials: S
authors.lastname: Mohri
mesh_terms: D000818:Animals; D051379:Mice; D008813:Mice, Inbred ICR; D018620:PrPSc Proteins / Q000737:chemistry / Q000378:metabolism*; D011328:Prions / Q000378:metabolism*; D017510:Protein Folding; D012608:Scrapie / Q000378:metabolism
chemical_list: D018620:PrPSc Proteins; D011328:Prions