[Establishment of PrP(Sc) conversion based on serial PMCA in vitro].
In order to establish an amplification system in vitro with which the PrP(Sc) is able to convert PrP(C) into proteinase K-resistant isoform infinitely and whether this system is more efficient than conventional protein misfolding cyclic amplification (PMCA), scrapie strain 263K-infected hamster's brain homogenate and homologous normal brain homogenate were prepared, respectively. A new methodology, namely serial PMCA, was utilized to reveal the continuous propagation ability of PrP(Sc). Totally 8 rounds of serial PMCA were proceeded and each round contained 48 cycles of alternative sonication and incubation. Simultaneously 144 cycles of conventional PMCA was used as a control. The results showed the PrP(Sc) from hamsters' brain tissues of scrapie agent 263K could replicate efficiently and infinitely with serial PMCA compared with finite propagation of PrP(Sc) with conventional PMCA system. The study of PrP(Sc) continuous propagation in brain homogenate with serial PMCA may further provide insight into the unsettled mechanism of prions misfolding and replication and apply to detect trace amount of PrP(Sc).
Shi
Song
S
State Key Laboratory for Infectious Disease Prevention and Control, National Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 100052, China. atfsu27@yahoo.com.cn
Dong
Chen-fang
CF
Zhang
Bao-yun
BY
Shi
Qi
Q
Wang
Gui-rong
GR
Wang
Xin
X
Han
Jun
J
Dong
Xiao-ping
XP
D000818:Animals; D001921:Brain / Q000378:metabolism; D006224:Cricetinae; D018621:PrPC Proteins / Q000737:chemistry; D018620:PrPSc Proteins / Q000737:chemistry*; D017510:Protein Folding
D018621:PrPC Proteins; D018620:PrPSc Proteins