Molecular specificities of antibodies against ovine and murine recombinant prion proteins.
The prion proteins (PrP) from sheep and mouse were produced in large quantities of full-length protein in Escherichia coli after fusion with a carboxy-terminal hexahistidine sequence. Both recombinant proteins were recognized, at variable levels, in ELISA using a panel of antibodies recognizing different parts of the PrP molecules, from the octo-repeat region (79-92 human sequence), to the C terminal end of the protein. We show that these recombinant proteins enable polyclonal antisera to be produced in PrP0/0 mice, the sheep prion protein being strongly immunogenic, using either native or guanidium hydrochloride-treated recombinant protein. Sera produced against the sheep protein also reacted in Western blot with bovine, ovine, and murine PrP res, but showed higher reactivity with sheep PrP res. Interestingly, when compared to an antiserum produced against bovine 106-121 peptidic sequence (RB1), we found strikingly different ratios of the PrP res glycoforms, in both cattle with BSE and sheep with natural scrapie, but not in scrapie infected mice. Such results further demonstrate that the assessment of PrP res glycoform ratios, using different antibodies, may depend on antibodies species-specificities.
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Agence Française de Sécurité Sanitaire des Aliments, Lyon, France.
Baron
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D000818:Animals; D000906:Antibodies; D015153:Blotting, Western; D001921:Brain / Q000378:metabolism; D002417:Cattle; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D019286:Endopeptidase K / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D018604:Epitope Mapping; D004926:Escherichia coli / Q000378:metabolism; D019791:Guanidine / Q000494:pharmacology; D006639:Histidine; D006801:Humans; D051379:Mice; D008957:Models, Genetic; D010455:Peptides / Q000737:chemistry / Q000378:metabolism; D011328:Prions / Q000276:immunology*; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000276:immunology; D011994:Recombinant Proteins / Q000276:immunology*; D012756:Sheep
D000906:Antibodies; D010455:Peptides; D011328:Prions; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; C033223:polyhistidine; D006639:Histidine; D019286:Endopeptidase K; D019791:Guanidine