Abstract
Transmissible spongiform encephalopathies (TSEs) are a group of invariably fatal neurodegenerative disorders. The causal agent is an aberrantly folded isoform (PrPSc or prion) of the endogenous prion protein (PrPC) which is neurotoxic and amyloidogenic and induces misfolding of its physiological counterpart. The intrinsic physical characteristics of these infectious proteinaceous pathogens makes them highly resistant to the vast majority of physicochemical decontamination procedures used typically for standard disinfection. This means prions are highly persistent in contaminated tissues, the environment (surfaces) and, of great concern, on medical and surgical instruments. Traditionally, decontamination procedures for prions are tested on natural isolates coming from the brain of infected individuals with an associated high heterogeneity resulting in highly variable results. Using our novel ability to produce highly infectious recombinant prions in vitro we adapted the system to enable recovery of infectious prions from contaminated materials. This method is easy to perform and, importantly, results in highly reproducible propagation in vitro. It exploits the adherence of infectious prion protein to beads of different materials allowing accurate and repeatable assessment of the efficacy of disinfectants of differing physicochemical natures to eliminate infectious prions. This method is technically easy, requires only a small shaker and a standard biochemical technique and could be performed in any laboratory.
Highlights
Transmissible spongiform encephalopathies (TSEs) or prion diseases are a group of invariably fatal neurodegenerative disorders that affect humans and a wide variety of mammals (Aguzzi and Calella, 2009)
All prion-coated beads made of different materials were able to give rise to Proteinase K-resistant misfolded recombinant PrP upon Protein Misfolding Shaking Amplification (PMSA) and showed the same electrophoretic mobility pattern and PK-resistant fragments (∼16, 9, and 2 kDa) as the original Sst01 (Figure 1)
The PMSA-derived products were digested with proteinase K (Roche) and subjected to electrophoresis and staining of total protein to assess the presence of proteinase K-resistant misfolded PrP in each sample
Summary
Transmissible spongiform encephalopathies (TSEs) or prion diseases are a group of invariably fatal neurodegenerative disorders that affect humans and a wide variety of mammals (Aguzzi and Calella, 2009). The main event in prion pathogenesis is the conformational conversion of the cellular prion protein (PrPC) into a pathological conformer (PrPSc) (Prusiner, 1982a) These two isoforms differ only in their three-dimensional structures and functionally in that the pathological isoform is both neurotoxic and able to induce conformation change of the normal cellular counterpart, PrPC, to the pathological isoform, PrPSc (Edgeworth et al, 2010). The latter, upon misfolding, contains a higher content of β-sheets and this makes it prone to aggregation, insolubility and partial
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