NR AOYS
AU Sarafoff,N.I.; Bieschke,J.G.; Giese,A.; Weber,P.; Piening,N.; Bertsch,U.; Kretzschmar,H.A.
TI Munich Prion Cycler (MPC) - an automated PMCA machine for therapeutic and diagnostic studies in prion diseases
QU International Conference - Prion diseases: from basic research to intervention concepts - TSE-Forum, 08.10.-10.10.2003, Gasteig, München - Poster session - IV-10
PT Konferenz-Poster
AB
Prions are the infectious agent of transmissible spongiforme encephalopathies (TSE) such as Creutzfeldt Jakob Disease and scrapie. Prions consist mainly of PrPsc, a proteinase K (PK)-resistant postranslationally misfolded isoform of the physiological copper binding prion protein PrPc. PrPsc autocatalytically induces the misfolding of PrPc, though the exact molecular mechanism of the conformational change still remains unknown. The Protein Misfolding Cyclic Amplification (PMCA) method is thought to model the conformational change in vitro. In a cyclic process consisting of sonication and incubation steps the physiological PrPc is changing its conformation to a PK resistant form (PrPres) in the presence of small amounts PrPsc. We characterized the sonication and incubation conditions for optimal amplification.
We showed that the amplification efficiency of the PK resistant PrPres may be increased by removing divalent cations with EDTA, whereas Cu, Mn, Zn and Ni ions decrease the PrPres amplification. The amplification may be achieved through direct as well as indirect sonication in a sonication bath. Indirect sonication allows the PMCA method to be automated. We developed a machine performing a PMCA reaction in a microplate format which promises to be a valuable tool in therapeutic and diagnostic screening of prion diseases.
AD N. Sarafoff, J. Bieschke, A. Giese, P. Weber, N. Piening, U. Bertsch, H.A. Kretzschmar, Institute for Neuropathology, LMU Munich, Marchioninistr. 17, 81377 Munich, Germany
SP englisch
PO Deutschland