Study for atypical BSE in Italy

2006 Fiscal Year Final Research Report Summary

• Project/Area Number: 17405042
• Research Category: Grant-in-Aid for Scientific Research (B)
• Allocation Type: Single-year Grants
• Section: 海外学術
• Research Field: Basic veterinary science/Basic zootechnical science
• Research Institution: The University of Tokyo
• Principal Investigator: ONODERA Takashi The University of Tokyo, Graduate School of Agricultural and Life Sciences, Professor, 大学院農学生命科学研究科, 教授 (90012781)
• Co-Investigator(Kenkyū-buntansha): SAEKI Keiichi The University of Tokyo, Graduate School of Agricultural and Life Sciences, Assistant, 大学院農学生命科学研究科, 助手 (10311630)
• Project Period (FY): 2005 – 2006
• Keywords: Italy / atypical / BSE / Western Blot / ELISA / monoclonal antibody / immunohistochemistry / bcl3 facilyty

Research Abstract

A monoclonal antibody (mAb) panel to bovine prion protein (PrP) was studied by immunoblotting and immunohistochemistry for scrapie, and bovine spongiform encephalopathy. The mAb panel is useful because they recognized both normal (PrP^c) and abnormal (PrP^<TSE>) isoforms of PrP in murine, ovine and bovine brain tissues. Among them, two mAbs recognized a conformational or fragmented PrP epitope. Interestingly, an anti-bovine PrP mAb, 1D12, prepared by immunizing PrP gene-knockout mice with a synthetic polypeptides corresponding to codons 153-166 of the bovine PrP gene showed the novel patterns of reactivity for prion-uninfected neuronal cells. When an enzyme-linked immunosorbent assay-mapping of the mAb epitopes was examined, monoclonal 1D12 reacted with YEDRY and M corresponding to amino acids 156-160 and 165 of bovine PrP. Several patterns of bovine PrP^c distribution in PrP-deficient neuronal cells (HpL3-4) transfected with bovine PrP were observed after different fixation methods. Staining of cell surface was observed after formalin fixation by immunofluorescent assay of 1D12 with confocal microscopy, whereas granules in nucleus were stained after acetone fixation. No reactivity in the nucleus was observed to HpL3-4 or HpL3-4 cells expressing mouse PrP. Reactivity in the nucleus was observed to HpL3-4 cells expressing hamster or bovine PrP. It is the first report to detect the PrPc at both cell surface and nuclei of prion-uninfected cell line. Furthermore, as nuclear PrP^c was specifically recognized by 1D12, amino acids 156-160 and 165 of hamster or bovine PrP may play important role in localization of PrP^c into nucleus.