Detection of prion particles in samples of BSE and scrapie by fluorescence correlation spectroscopy without proteinase K digestion.
A characteristic feature of prion diseases such as bovine spongiform encephalopathy (BSE) is the accumulation of a pathological isoform of the host-encoded prion protein, PrP. In contrast to its cellular isoform PrP(C), the pathological isoform PrP(Sc) forms insoluble aggregates. All commercial BSE tests currently used for routine testing are based on the proteinase K (PK) resistance of PrP, but not all pathological PrP is PK-resistant. In the present study, single prion particles were counted by fluorescence correlation spectroscopy (FCS). The property of PK resistance is not required, i.e., both the PK-resistant and the PK-sensitive parts of the prion particles are detectable. PrP aggregates were prepared from the brains of BSE-infected cattle, as well as from scrapie-infected hamsters, by the NaPTA precipitation method without PK digestion. They were labeled using two different PrP-specific antibodies for FCS measurements in the dual-color mode (2D-FIDA). Within the limited number of samples tested, BSE-infected cattle and scrapie-infected hamsters in the clinical stage of the disease could be distinguished with 100% specificity from a control group. Thus, a diagnostic tool for BSE detection with complete avoidance of PK treatment is presented, which should have particular advantages for testing animals in the preclinical stage.
Heinrich-Heine-Universität Düsseldorf, Institut für Physikalische Biologie, Universitätsstr. 1, D-40225 Düsseldorf, Germany.
Eva
E
Birkmann
Oliver
O
Schäfer
Nicole
N
Weinmann
Christian
C
Dumpitak
Michael
M
Beekes
Roy
R
Jackman
Leigh
L
Thorne
Detlev
D
Riesner
D000818:Animals; D002417:Cattle; D006224:Cricetinae; D016643:Encephalopathy, Bovine Spongiform / Q000175:diagnosis*; D019286:Endopeptidase K / Q000737:chemistry*; D008526:Medulla Oblongata / Q000737:chemistry; D011328:Prions / Q000302:isolation & purification*; D012608:Scrapie / Q000175:diagnosis*; D013050:Spectrometry, Fluorescence / Q000379:methods
D011328:Prions; D019286:Endopeptidase K