Aguzzi
Fakultäten » Medizinische Fakultät » Neuropathologie, Institut für » Prof. Dr. Adriano Aguzzi » Aguzzi
Completed research project
| Title / Titel | Application of the Transient Scrapie Cell Assay (TraSCA) for in vitro detection of ovine and bovine prions | ||
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| Abstract (PDF, 14 KB) | |||
| Summary / Zusammenfassung | Detection of prion infectivity in animal tissues and fluids traditionally relies on murine bioassays (MBA) and thus is significantly time consuming, expensive and involves the use of large numbers of mice. An in vitro assay (Scrapie Cell Assay “SCA”) for determination of prion infectivity, as sensitive as the MBA has recently been developed. However, the assay is still time consuming and detection is so far restricted to mouse-adapted scrapie strains. At present we are in the process of successfully improving the SCA as originally published and developing of a murine Transient Scrapie Cell Assay (TraSCA) for tagged versions of murine PrPs. The advantage of TraSCA technology is the circumvention of time consuming passaging procedures, therefore allowing for dramatic shortening in assay time. We propose to extend the applicability of the TraSCA and to develop the ovine and bovine TraSCA for the rapid measurement of prion infectivity in ovine and bovine tissues and fluids. Epitope-tagged PrPC allows detection of de novo formed PrPSc within 24 hours post exposure to prions. We will therefore develop a modification of the SCA, forthwith denominated Transient Scrapie Cell Assay (TraSCA), utilising murine and ovine cell lines stably expressing tagged ovine and bovine PrPs. Discrimination of tagged PrPSc from tagged PrPC requires a proteinase K digest step in the TraSCA. To avoid this we also intend to investigate the feasibility of utilising tetracysteine tags and the FlAsH technology in PrP. Success of such technology may allow for high-throughput fluoresence activated cell sorting (FACS)-based platforms to be used for readout of the TraSCA. It is anticipated that this project will allow for the use of the TraSCA to determine prion infectivity in tissue homogenates and fluids of sheep and cows. A cell-based infectivity assay such as the TraSCA for ovine and bovine prions will allow for sensitive high throughput screening of risk material. In addition, TraSCA technology will not only dramatically reduce the number of mice used in prion research but also significantly accelerate experiments and analysis of prion infectivity in ovine and bovine tissues and fluids, thereby greatly advancing research on TSEs. |
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| Keywords / Suchbegriffe | TSE, BSE, scrapie, prion infectivity, bioassay | ||
| Project leadership and contacts / Projektleitung und Kontakte |
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| Funding source(s) / Unterstützt durch |
Others DEFRA, UK |
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| Duration of Project / Projektdauer | Sep 2005 to Aug 2008 |