NR AXPI
AU Jones,M.; Head,M.W.; Connolly,J.G.; Farquhar,C.F.; MacGregor,I.R.
TI Novel Application Specific Monoclonal Antibodies Targeting Human Prion Protein
QU International Conference - Prion 2007 (26.-28.9.2007) Edinburgh International Conference Centre, Edinburgh, Scotland, UK - Book of Abstracts: Protein Misfolding P01.48
IA http://www.prion2007.com/pdf/Prion Book of Abstracts.pdf
PT Konferenz-Poster
AB
Background: Monoclonal antibodies (mAbs) raised against human prion protein (PrP) may prove to be valuable reagents in the detection, treatment and general understanding of human prion diseases, however to date no such antibodies are available.
Aim(s)/Objectives(s): To develop a method for the purification, under native conditions, of cellular prion protein (PrPc) from human platelets and to use this material as an immunogen to raise a panel of mAbs targeting human PrP.
Methods: PrPc was purified from human platelet lysates using a combination of cation exchange and Cu2+ affinity chromatography. This purified PrPc was used to immunize PrP null mice, spleen cells from the immunised mice were fused to SP2/0 myeloma cells and stable hybridomas, secreting anti-PrP mAbs, were single cell cloned. The mAbs produced were characterized by isotype; epitope mapping; binding to both native/denatured platelet PrP and native/denatured a-helical/ß-sheet recombinant mouse PrP; immunoprecipitation of PrPc, disease associated PrP (PrPsc) and its corresponding proteinase K resistant core (PrP27-30) from neurological control/vCJD brain homogenates.
Results: Following immunization with purified platelet PrPc, all mice developed a strong, predominantly IgG isotype, anti-PrP immune response. From four separate fusions a total of twelve stable hybridoma cell lines secreting anti-HuPrP mAbs were identified and single cell cloned. A number of these mAbs displayed novel PrP binding properties, cross-reactivity with mouse PrP, differential binding to native/denatured PrP and PrPsc/PrP27-30 selectivity in immunoprecipitation experiments.
Conclusions: A panel of 12 mAbs raised against native human platelet PrPc has been produced. Initial characterisation suggests that some of these antibodies may prove to be useful reagents not only in the detection, treatment and understanding of human prion diseases but in TSE research more generally.
AD M. Jones, M.W. Head, University of Edinburgh, National CJD Surveillance Unit, UK; J.G. Connolly, University of Strathclyde, SIPBS, UK; C.F. Farquhar, Roslin Institute Neuropathogenesis Unit, UK; I.R. MacGregor, Scottish National Blood Transfusion Service, National Science Laboratory, UK
SP englisch
PO Schottland