A direct assessment of human prion adhered to steel wire using real-time quaking-induced conversion.

Tsuyoshi Mori,Takehiro Nakagaki,Masahisa Hamada,Kazunori Sano,Takehisa Nakayama,Kazuko Ichimiya,Daisuke Ishibashi,Hanae Takatsuki,Noriyuki Nishida,Ryuichiro Atarashi,Katsuya Satoh,Kana Furukawa

doi:10.1038/srep24993

Abstract

Accidental transmission of prions during neurosurgery has been reported as a consequence of re-using contaminated surgical instruments. Several decontamination methods have been studied using the 263K-hamster prion; however, no studies have directly evaluated human prions. A newly developed in vitro amplification system, designated real-time quaking-induced conversion (RT-QuIC), has allowed the activity of abnormal prion proteins to be assessed within a few days. RT-QuIC using human recombinant prion protein (PrP) showed high sensitivity for prions as the detection limit of our assay was estimated as 0.12 fg of active prions. We applied this method to detect human prion activity on stainless steel wire. When we put wires contaminated with human Creutzfeldt–Jakob disease brain tissue directly into the test tube, typical PrP-amyloid formation was observed within 48 hours, and we could detect the activity of prions at 50% seeding dose on the wire from 102.8 to 105.8 SD50. Using this method, we also confirmed that the seeding activities on the wire were removed following treatment with NaOH. As seeding activity closely correlated with the infectivity of prions using the bioassay, this wire-QuIC assay will be useful for the direct evaluation of decontamination methods for human prions.

Highlights

Infectious agents are highly resistant to routine decontamination methods[13]
The QuIC signal could detect prion seeds attached to the wire (Fig. 1)
NaOH treatment removed the positive signal of real-time quaking-induced conversion (RT-QuIC), there was no significant change following incubation with Sodium dodecyl sulfate (SDS) solution

Summary

Introduction

Infectious agents are highly resistant to routine decontamination methods[13]. High concentrations of sodium hydroxide, sodium hypochlorite or prolonged steam sterilisation are recommended methods for prion disinfection; most methods damage the surgical instruments[14,15,16,17]. Because of the lack of nucleic acid components, approaches for TSE rely upon methods of immunodetection including immunohistochemistry and enzyme-linked immunosorbent assay using antibodies against PrP18–21. Another evaluation method is Western blotting for protease-resistant PrP22. We demonstrated that treatment of wire with 1 mol/L NaOH solution was suitable for decontamination of prions. These results indicate that wire-QuIC can be useful to evaluate the decontamination of human prions on medical devices such as surgical instruments

Methods

Results

Conclusion