NR ATKB
AU Leclere,E.; Lavigne,C.; Frobert,E.; Calavas,D.; Baron,T.G.M.; Perron,H.; Moussa,A.
TI PrPres precipitation by Streptomycin is more sensitive than ultracentrifugation for brain and spleen WB analysis in animal transmissible spongiform encephalopathies
QU International Conference - Prion 2005: Between fundamentals and society's needs - 19.10.-21.10.2005, Congress Center Düsseldorf - Poster Session: Diagnosis DIA-13
PT Konferenz-Poster
AB
Transmissible spongiform encephalopathies or prion diseases, are a group of mammalian neurodegenerative disorders characterized by a posttranslational conformational modification and brain accumulation of an insoluble protease-resistant isoform of the host encoded cellular prion protein. At present, diagnostic confirmation of the bovine disease, bovine spongiform encephalopathy, is based on Western blot assay with ultracentrifugation. In order to achieve a concentration step compatible with routine diagnostic procedures, we have investigated various molecules for their interaction with PrPsc. One molecule when used certain conditions, Streptomycin, was found to aggregate PrPres fragment remaining in BSE and scrapie brain samples after PK digestion.
We have compared the effect of Streptomycin and ultracentrifugation for the concentration and isolation of PrPres in 10% homogenates (w/v) of brains and spleens from different species. After PK digestion, samples were ultracentrifugated at 465,000Xg for 2 hours or precipitated with Streptomycin and centrifuged at 12,000 for 5 minutes. Pellets were resuspended in 50µl of denaturing buffer before Western blot analysis and SAF84 monoclonal specific antibody detection.
The results indicate that, with the same amount of sample tested, Streptomycin permits a more sensitive detection of PrPres than ultracentrifugation.
Therefore, streptomycine provides a technical avenue for a rapid diagnosis confirmation for BSE as well as ovine or murine Scrapie in brain or spleen, but may also be extended to other organs or tissues.
IN Die Autoren erreichten durch eine Fällung mit Streptomycin eine höhere Konzentration der PrPres als durch die übliche Ultrazentrifugation.
AD E.Leclere, H.Perron, bioMérieux R&D immunoassay and Proteomics dept, Marcy L'Etoile, France.; C.Lavigne, E.Frobert, D.Calavas, T.Baron, A.Moussa, Agence Française pour la Sécurité Sanitaire des Aliments, Lyon, France.
SP englisch
PO Deutschland