From a prion seed to a tree of discoveries: Byron Caughey's RT-QuIC legacy in protein misfolding research.

Early and accurate detection of prions is essential for effective disease management and potential therapeutic intervention. Similar challenges extend across other protein misfolding diseases, which require ultrasensitive methods for detection, discrimination, and quantification of "prion-like" pathological proteins. Over a scientific career spanning multiple decades, Dr. Byron Caughey investigated mechanisms of prion conversion and replication, pioneered cell-free prion conversion assays, and steadfastly pursued increasingly sensitive, specific, robust, and high-throughput methods for prion amplification. His breakthrough development and continued refinement of the Real-Time Quaking-Induced Conversion (RT-QuIC) assay revolutionized prion research and helped extend seed amplification technology across the broader field of protein misfolding diseases. These assays have been developed through seminal contributions by many eminent scientists, and the breadth of this rapidly expanding field extends well beyond the scope of this review. Accordingly, we focus here on honoring Dr. Byron Caughey's legacy by highlighting his foundational contributions to in vitro amplification and ultrasensitive detection of disease-associated protein aggregates. We further highlight the visionary, multidisciplinary, and collaborative nature of his work including the expansion of the platform to strain discrimination, surveillance, misfolded protein quantification, therapeutic research, higher throughput assay formats and the detection of other disease-associated misfolded proteins, including α-synuclein, tau, SOD1, and more.