Sensitive detection of pathological seeds of α-synuclein, tau and prion protein on solid surfaces.

Prions or prion-like aggregates such as those composed of PrP, α-synuclein, and tau are key features of proteinopathies such as prion, Parkinson's and Alzheimer's diseases, respectively. Their presence on solid surfaces may be biohazardous under some circumstances. PrP prions bound to solids are detectable by ultrasensitive real-time quaking-induced conversion (RT-QuIC) assays if the solids can be immersed in assay wells or transferred to pads. Here we show that PrP prions can remain detectable on steel wires for at least a year, or even after enzymatic cleaning and sterilization. We also show that contamination of larger objects with pathological seeds of α-synuclein, tau, and PrP can be detected by simply assaying a sampling medium that has been transiently applied to the surface. Human α-synuclein seeds in dementia with Lewy bodies brain tissue was detected by α-synuclein RT-QuIC after drying of tissue dilutions with concentrations as low as 10⁻⁶ onto stainless steel. Tau RT-QuIC detected tau seeding activity on steel exposed to Alzheimer's disease brain tissue diluted as much as a billion fold. Prion RT-QuIC assays detected seeding activity on plates exposed to brain dilutions as extreme as 10⁻⁵–10⁻⁸ from prion-affected humans, sheep, cattle and cervids. Sampling medium collected from surgical instruments used in necropsies of sporadic Creutzfeldt-Jakob disease-infected transgenic mice was positive down to 10⁻⁶ dilution. Sensitivity for prion detection was not sacrificed by omitting the recombinant PrP substrate from the sampling medium during its application to a surface and subsequent storage as long as the substrate was added prior to performing the assay reaction. Our findings demonstrate practical prototypic surface RT-QuIC protocols for the highly sensitive detection of pathologic seeds of α-synuclein, tau, and PrP on solid objects. Author summary: Prions and prion-like protein aggregates associated with neurodegenerative diseases such as Creutzfeldt-Jakob disease, Alzheimer's disease, and dementia with Lewy bodies may persist in the environment or on medical instruments and in some cases be transmissible. Sensitive detection of these self-propagating aggregates, or seeds, on solid surfaces, machinery, tools, and surgical instruments might help prevent fomite-borne disease dissemination. Here we describe a novel RT-QuIC (sfRT-QuIC) to detect a variety of pathological brain-derived prion, α-synuclein, or tau seeds on stainless steel and acrylic surfaces. sfRT-QuIC testing showed that α-synuclein and tau seeds remained active on surgical forceps and scissors, even after routine instrument sterilization. Furthermore, we found that pathological protein seeds eluted from surfaces could be collected and stored for transport or delayed testing. sfRT-QuIC is a simple and effective, high throughput assay for sensitive detection of pathological protein seed contamination of common surfaces. [ABSTRACT FROM AUTHOR]