Detection of bovine spongiform encephalopathy, ovine scrapie prion-related protein (PrPSc) and normal PrPc by monoclonal antibodies raised to copper-refolded prion protein.
Prion-related protein (PrP) is a glycosylphosphatidylinositol-linked cell-surface protein expressed by a wide variety of cells, including those of the nervous system and the immune system. Several functions of normal cellular PrP (PrPc) have been proposed that may be associated with the capacity of this protein to bind copper. In the present study, we describe the generation of a panel of monoclonal antibodies raised to copper-refolded PrP, which may be used to analyse the normal and disease-associated forms of this protein. The anti-PrP monoclonal antibodies were reactive by Western blot and ELISA with recombinant murine PrPc refolded in the presence or absence of either copper or manganese, and with the disease-susceptible allelic form V136R154Q171 ('VRQ'; where single-letter amino-acid notation has been used) and disease-resistant allelic form A136R154R171 ('ARR') of recombinant ovine PrPc. FACS analysis of lymphoid cells using these monoclonal antibodies showed that wild-type non-activated mouse lymphocytes expressed little, if any, PrPc. These monoclonal antibodies were shown to react with the unglycosylated and monoglycosylated forms of PrPSc (abnormal disease-specific conformation of PrP) in prion-infected tissue samples from all of the different species tested by Western blot. In addition, this analysis allowed one to make a distinction between bovine spongiform encephalopathy ('BSE') and scrapie PrPSc) isolates from experimentally infected sheep on the basis of their different electrophoretic mobilities.
Thackray
Alana M
AM
Centre for Veterinary Science, Department of Clinical Veterinary Medicine, University of Cambridge, Madingley Road, Cambridge CB3 0ES, UK.
Madec
Jean-Yves
JY
Wong
Edmond
E
Morgan-Warren
Robert
R
Brown
David R
DR
Baron
Thierry
T
Bujdoso
Raymond
R
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D015153:Blotting, Western; D002417:Cattle; D003300:Copper / Q000378:metabolism*; D003429:Cross Reactions; D016643:Encephalopathy, Bovine Spongiform / Q000175:diagnosis*; D004797:Enzyme-Linked Immunosorbent Assay; D018604:Epitope Mapping; D005434:Flow Cytometry; D006031:Glycosylation; D010446:Peptide Fragments / Q000302:isolation & purification; D018621:PrPC Proteins / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification*; D018620:PrPSc Proteins / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification*; D017510:Protein Folding; D012756:Sheep
D000911:Antibodies, Monoclonal; D010446:Peptide Fragments; D018621:PrPC Proteins; D018620:PrPSc Proteins; D003300:Copper