Role of different recombinant PrP substrates in the diagnostic accuracy of the CSF RT-QuIC assay in Creutzfeldt-Jakob disease

Abstract

The development of the real-time quaking-induced conversion (RT-QuIC), an in vitro protein misfolding amplification assay, was an innovation in the scientific field of protein misfolding diseases. In prion diseases, these types of assays imitate the pathological conversion of the cellular prion protein (PrPC) into a protease-resistant and/or amyloid form of PrP, called PrP resistant (PrPRes). The RT-QuIC is an automatic assay system based on real-time measuring of thioflavin-T (Th-T) incorporation into amyloid fibrils using shaking for disaggregation. It has already been applied in diagnostics, drug pre-screening, and to distinguish between different prion strains. The seeded conversion efficiency and the diagnostic accuracy of the RT-QuIC assay strongly depend on the kind of recombinant PrP (rec PrP) substrate. The DNA sequences of different substrates may originate from different species, such as human, bank vole, and hamster, or from a combination of two species, e.g., hamster-sheep chimera. In routine use, either full-length (FL) or truncated substrates are applied which can accelerate the conversion reaction, e.g., to a more sensitive version of RT-QuIC assay. In the present review, we provide an overview on the different types of PrP substrates (FL and truncated forms), recapitulate the production and purification process of different rec PrP substrates, and discuss the diagnostic value of CSF RT-QuIC in human prion disease diagnostics.

New developments in prion disease research

“…Ryuichiro Atarashi describes the technology and provides an overview of how it is already used in diagnostics and what the potential for pre-clinical diagnosis and treatment monitoring is (Atarashi 2022). Inga Zerr and colleagues discuss how the diagnostic accuracy of the RT-QuIC assay depends on the recombinant PrP used as substrate, focusing on the diagnostic value of CSF RT-QuIC in the diagnostics of human prion disease (da Silva Correia et al 2022). Protein misfolding cyclic amplification, or short PMCA, was the first ultrasensitive prion amplification method to be established.…”