Masuhiro Takata

Rapid PrPSc Detection in Lymphoid Tissue and Application to Scrapie Surveillance of Fallen Stock in Japan: Variable PrPSc Accumulation in Palatal Tonsil in Natural Scrapie

Rapid western blot (WB) procedure for an abnormal isoform of prion protein (PrPSc) detection in lymphoid tissues was established and has been applied to the surveillance of fallen stock. In this program, brain and palatal tonsil were examined by WB and three cases of sheep scrapie were detected. While one clinically scrapie‐infected sheep harbored PrPSc in the brain and palatal tonsil, the two sheep in the pre‐clinical stage harbored PrPSc in the brain, but not in the palatal tonsil. This study shows that PrPSc accumulation in palatal tonsil is variable in natural scrapie, even among genetically susceptible sheep.

A Potential Blood Test for Transmissible Spongiform Encephalopathies by Detecting Carbohydrate-Dependent Aggregates of PrPres-Like Proteins in Scrapie-Infected Hamster Plasma

PrPres has rarely been detected in blood (except in leukocytes) even in diseased animal models that are known to contain a large amount of PrPres in infected tissues. It seems likely that PrPres detection in blood is difficult because of the low titer of infectious material within the blood. Here, we demonstrate the detection of proteinase K‐resistant 3F4‐reactive protein in the plasma of scrapie‐infected hamsters but not in the plasma of mock‐infected hamsters by partial purification using a novel method termed “acidic SDS precipitation,” in conjunction with a highly sensitive chemiluminescence detection system used to show the presence of PrP at a concentration equivalent to 1.4 ×10–9 g of brain homogenate or 1.5 × 10–12 g (6.5 ×10–17 mol) of rPrP by conventional Western blotting. The 3F4‐reactive proteins in scrapie‐infected hamster plasma often resulted in multiple Mw protein bands occurring at higher Mw positions than the position of the di‐glycosyl PrP molecule. Mixing scrapie‐infected hamster brain homogenate with mock‐infected hamster plasma resulted in the formation of similar Mw positions for multiple 3F4‐reactive proteins. Predigestion of carbohydrate side chains from the proteins in the plasma or brain homogenate before mixing resulted in failure to obtain these multiple 3F4‐reactive proteins. These observations indicate that PrPres aggregated with other proteins in the plasma through carbohydrate side chains and was successfully detected in the plasma of scrapie‐infected hamsters. Counterparts in these aggregates with PrPres‐like proteins in scHaPl are not known but any that exist should resist the PK digestion.