Efficient in vitro amplification of a mouse-adapted scrapie prion protein

Protein misfolding cyclic amplification (PMCA) is a highly sensitive technique used to detect minute amounts of scrapie prion protein (PrPSc), a major protein component of the infectious agents associated with prion diseases. Although exponential in vitro amplification of hamster scrapie PrPSc has been established, the PMCA used was unsuccessful in achieving good amplification of PrPSc from other animals. Here, we have investigated the cause of the insufficient PrPSc amplification in mice and have developed an improved method suitable for amplification of the PrPSc of the mouse-adapted scrapie prion strain Chandler. Mouse PrPC, 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 PrPC was inhibited; hence, mouse PrPSc could be amplified to infinite levels. The present study is the first report describing effective amplification of PrPSc of the mouse-adapted scrapie prion and this improved PMCA technique will contribute to prion research that uses mice as experimental animals.