NR AWHZ
AU König,C.; Engemann,C.; Krummrei,U.; Wacker,R.; Hoffmann,R.
TI Detection of prions in body fluids - diagnosis of CJD
QU International Conference - Prion 2006: Strategies, advances and trends towards protection of society - 3.10.-6.10.2006, Torino, Italy, Lingotto Conference Centre - Poster sessions DIA-30
PT Konferenz-Poster
AB Transmissible spongiform encephalopathies (TSEs) are fatal disorders of the central nervous system characterised by the progressive accumulation of an abnormal form of the prion protein (PrP). TSEs include scrapie in sheep, bovine spongiform encephalopathy (BSE) in cattle, and Creutzfeldt-Jakob disease (CJD) in humans. It was shown that BSE can be transmitted by the consumption of meat products from BSE infected cattle to humans causing the new variant of CJD (nvCJD) in humans. Among humans nvCJD can be transmitted by blood transfusions and organ transplantations. Due to the slow etiopathology, a general screening of all donors appears obligatory, as the infection risk of a transplant can be unperceived. Thus, our ambition was the development of a detection system for PrPsc in body fluids. At present, there are no methods available that allow the detection of PrPsc in body fluids, because of the extremely low concentrations expected. Therefore, a substantial increase in sensitivity of prion detection systems is necessary. On the basis of a classical ELISA we could establish such a highly sensitive test system by using a new detection technique, the so called Immuno-PCR. The ImmunoPCR combines the specific antigen-antibody reaction with the high amplification rate in PCR. Using Immuno-PCR, sensitivity of the corresponding classical ELISA was increased 10.000 fold resulting in a detection limit of 10pg/mL recombinant prion protein. By spiking experiments using recombinant prion protein in body fluids it was shown that 10pg/mL recPrP are clearly detectable even when spiked in serum or cerebrospinal fluid. In a further experiment brain homogenate from a BSE-infected macaque and a healthy animal were analyzed in Immuno-PCR. The results demonstrate that PrPsc from animals which were infected with BSEpositive material can be detected with the presented technique. Next, the preparation of serum and cerebrospinal fluid was optimized in terms of buffer composition and Proteinase K treatment. Finally, samples from patients with various neurodegenerative diseases were analyzed by the optimized sample preparation combined with the Immuno-PCR. First results on the quantification of PrPsc in human body fluid samples from patients compared to healthy control individuals will be presented.
AD C. König, R. Hoffmann: Biotechnologisch-Biomedizinisches Zentrum, AG Bioanalytik, Universität Leipzig, Germany; C. Engemann, U. Krummrei: Priontype GmbH & Co.KG, Leipzig, Germany; R. Wacker: Chimera Biotec GmbH, Dortmund, Germany
SP englisch
PO Italien