Western Blot Detection of Scrapie-associated Fibril Protein in Tissues outside the Central Nervous System from Preclinical Scrapie-infected Mice

Abstract

SUMMARYWe describe a method of sample preparation to detect scrapie-associated fibril (SAF) proteins in small amounts of scrapie-infected mouse tissues by Western blot analysis using an antiserum to a synthetic peptide that corresponds to the N-terminal region of hamster prion protein. SAF proteins were efficiently detected in brain tissue by this procedure. The proteins were also detected in preparations from spleen and lymph node. SAF proteins were detected in brain samples at 24 weeks after intraperitoneal infection. Using spleen samples, the proteins were detected from mice in the preclinical stage (from 4 weeks after infection), clinical symptoms of scrapie were observed in some mice from 22 weeks after infection.

Detection of proteinase K-resistant prion protein and infectivity in mouse spleen by 2 weeks after scrapie agent inoculation

“…), which again is well after infectivity levels in spleen plateau. However, Doi et al (1988) were able to detect PrP-res in mouse spleen by 5 weeks p.i., making the association with infectivity more likely, but they did not determine infectivity. Considered together, these data show wide variation with regard to the time p.i.…”

“…Their spleens and brains were harvested aseptically and frozen until they could be analysed. PrP-res preparations were made from pooled spleens using several modifications of a technique described previously (Doi et al, 1988). Briefly, 10% suspensions of spleen in 0.01 M-Tris-HC1, 0.005 M-MgC12 were made by forcing the spleens through a fine mesh stainless steel screen.…”

Strain specific and common pathogenic events in murine models of scrapie and bovine spongiform encephalopathy

“…hypertrophy and possible proliferation of astrocytes (Fraser, 1976). The most specific molecular feature is the accumulation in the brain (Prusiner, 1982) and some peripheral organs (Doi et al, 1988;Farquhar et al, 1994) of an abnormal isoform of the host-encoded PrP protein, which copurifies with infectivity (Bolton et al, 1982;McKinley et al, 1983). The pathological isoform has been named PrP se for scrapie, or PrP res for its enhanced resistance to proteases (Bolton et al, 1982;Prusiner et al, 1983).…”

Feline spongiform encephalopathy

“…This technique has been used to identify experimentally induced infections of scrapie in both mice and sheep before clinical signs develop (Doi andothers 1988, Ikegami andothers 1991). However, replication of TSE agents in lymphoreticular tissue has been shown to be much more limited in certain of the TSE conditions such as in TME (Hadlow and others 1987).…”