Applicability of three anti-PrP peptide sera including staining of tonsils and brainstem of sheep with scrapie.
Three rabbit antibodies (R521, R505, R524) were produced, and raised to synthetic peptides corresponding to residues 94-105, 100-111, and 223-234, respectively, of the sheep prion protein (PrP). Epitope mapping analysis revealed the monospecific character of antisera R505 and R524. In addition to the amino acid sequence against which it was raised, R521 also recognized other small epitopes. ELISA and radio-immunoprecipitation were used to assess the relative immunoreactivities of the antisera to the normal sheep prion protein (PrP(c)). Highest reactivity was found for R521, followed by R505 and R524. According to Western blot analysis, all three sera specifically reacted with the prion proteins PrP(Sc) and PrP27-30, extracted from the brain stem of a scrapie-affected sheep. Yet, with R505 not all of the lower molecular weight deglycosylated forms could be detected. Contrary to the immunoreactivities found with the PrP(Sc) and PrP27-30 isoforms, only R521 recognised PrP(c) from a healthy sheep. The usefulness of all three anti-peptide sera in the immunohistochemical detection of PrP(Sc) in brain stem and tonsils of scrapie-affected sheep was demonstrated and compared with an established rabbit anti-PrP serum.
Garssen
G J
GJ
Department of Molecular Recognition, Institute for Animal Science and Health (ID-Lelystad), Lelystad, The Netherlands. g.j.garssen@id.wag_ur.nl
Van Keulen
L J
LJ
Farquhar
C F
CF
Smits
M A
MA
Jacobs
J G
JG
Bossers
A
A
Meloen
R H
RH
Langeveld
J P
JP
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000032:analysis; D000918:Antibody Specificity; D015153:Blotting, Western; D001921:Brain / Q000378:metabolism* / Q000473:pathology; D004797:Enzyme-Linked Immunosorbent Assay; D018604:Epitope Mapping; D007106:Immune Sera / Q000032:analysis; D007150:Immunohistochemistry / Q000379:methods; D051379:Mice; D008969:Molecular Sequence Data; D014066:Palatine Tonsil / Q000378:metabolism* / Q000473:pathology; D010455:Peptides / Q000138:chemical synthesis / Q000276:immunology; D018621:PrPC Proteins / Q000276:immunology; D018620:PrPSc Proteins / Q000276:immunology; D011233:Precipitin Tests; D011328:Prions / Q000032:analysis* / Q000276:immunology; D020033:Protein Isoforms / Q000276:immunology; D011994:Recombinant Proteins / Q000276:immunology; D012608:Scrapie / Q000378:metabolism* / Q000473:pathology; D012756:Sheep; D014407:Tumor Cells, Cultured
D000906:Antibodies; D007106:Immune Sera; D010455:Peptides; D018621:PrPC Proteins; D018620:PrPSc Proteins; D011328:Prions; D020033:Protein Isoforms; D011994:Recombinant Proteins