NR AXOJ
AU Henke,F.; Funke,A.; Willbold,D.; Riesner,D.; Birkmann,E.
TI Detection of Single Scrapie- and Bse-Prion Particles by Surface-Fida
QU International Conference - Prion 2007 (26.-28.9.2007) Edinburgh International Conference Centre, Edinburgh, Scotland, UK - Book of Abstracts: Epidemiology, Risk Assessment and Transmission P04.106
IA http://www.prion2007.com/pdf/Prion Book of Abstracts.pdf
PT Konferenz-Poster
AB
The infectious agents of prion diseases are composed primarily of the pathogenic isoform of the prion protein designated PrPsc, which is generated by a conformational change of the cellular isoform PrPc. In contrast to its cellular isoform, the pathogenic isoform PrPsc forms insoluble aggregates. Hitherto approved prion tests use the PK-resistance of PrPsc as a marker for the disease. Because of varying portions of disease related aggregated PrP, which is not PK-resistant [2; 3], these prion tests offer only a limited sensitivity.
We developed a new method for prion detection with two major advantages. First it avoids completely PK-digestion, and second it counts single prion particles. The detection system is based on Fluorescence-Correlation-Spectroscopy (FCS). The partially purified prion particles are labelled by two different antibodies with different fluorescence labels, so that FCS is applied in the mode of Dual-Colour FluorescenceIntensity-
Distribution-Analysis (2D-FIDA) [2]. To increase the sensitivity particles were concentrated on a chip surface by capture antibodies (Surface-FIDA) [1].
With Surface-FIDA we are able to distinguish Scrapie-infected hamster as well as BSEinfected cattle in the clinical stage from a control group [1]. Therefore the sensitivity to identify Scrapie-infected hamster as well as BSE infected cattle in brain samples could be increased dramatically as compared to FIDA in solution [2]. Preliminary data showed that applying Surface-FIDA one is able to detect PrP-aggregates in the cerebrospinal fluid of cattle. We present further optimization of Surface-FIDA in respect of the methodology to enhance specificity and sensitivity.
[1] Birkmann et al., Counting of single prion particles bound to a capture-antibody surface (Surface-FIDA), J. Vet. Microbiol. (in press) [2] Birkmann et al., (2006) Detection of prion particles in samples of BSE and Scrapie by Fluorescence Correlation Spectroscopy without Proteinase K digestion. Biol Chem., 387, 95-102. [3] Safar, J. et al. (1998). Eight prion strains have PrPsc molecules with different conformations. Nat Med. 10, 1157-1165.
AD F. Henke, A. Funke, D. Willbold, D. Riesner, E. Birkmann, Heinrich-Heine-Universität, Institut für Physikalische Biologie, Germany
SP englisch
PO Schottland