A 2011 study reported the detection of prions in urine from naturally and orally infected sheep with clinical scrapie agent and orally infected preclinical and infected white-tailed deer with clinical chronic wasting disease (CWD). This is the first report on prion detection of PrP from the urine of naturally or preclinical prion-diseased ovines or cervids.
A 2010 study demonstrated a moderate amount of protein misfolding cyclic amplification (PMCA) coupled to a novel SOFIA detection scheme, can be used to detect PrP in protease-untreated plasma from preclinical and clinical scrapie sheep, and white-tailed deer with chronic wasting disease, following natural and experimental infection. The disease-associated form of the prion protein (PrP), resulting from a conformational change of the normal (cellular) form of prion protein (PrP), is considered central to neuropathogenesis and serves as the only reliable molecular marker for prion disease diagnosis. While the highest levels of PrP are present in the CNS, the development of a reasonable diagnostic assay requires the use of body fluids which characteristically contains extremely low levels of PrP. PrP has been detected in the blood of sick animals by means of PMCA technology. However, repeated cycling over several days, which is necessary for PMCA of blood material, has been reported to result in decreased specificity (false positives). To generate an assay for PrP in blood that is both highly sensitive and specific, the researchers used limited serial PMCA (sPMCA) with SOFIA. They did not find any enhancement of sPMCA with the addition of polyadenylic acid, nor was it necessary to match the genotypes of the PrP and PrP sources for efficient amplification.
A 2009 study found SOFIA, in its current format, is capable of detecting less than 10 attogram (ag) of hamster, sheep and deer recombinant PrP. About 10 ag of PrP from 263K-infected hamster brains can be detected with similar lower limits of PrP detection from the brains of scrapie-infected sheep and deer infected with chronic wasting disease. These detection limits allow protease-treated and untreated material to be diluted beyond the point where PrP, nonspecific proteins or other extraneous material may interfere with PrP signal detection and/or specificity. This not only eliminates the issue of specificity of PrP detection, but also increases sensitivity, since the possibility of partial PrP proteolysis is no longer a concern. SOFIA will likely lead to early ante mortem detection of transmissible encephalopathies and is also amenable for use with additional target amplification protocols. SOFIA represents a sensitive means for detecting specific proteins involved in disease pathogenesis and/or diagnosis that extends beyond the scope of the transmissible spongiform encephalopathies.