Method of the Month — RT-QuIC
May 2021
Our method for this month is Real-time Quaking Induced Conversion (RT-QuIC). This is a test that can detect prion proteins and is used clinically to diagnose Creutzfeldt-Jakob disease in humans.
RT-QuIC is a test used to detect diseased prion proteins (PrPSc), which are the infectious agent of prion diseases like Bovine Spongiform Encephalopathy, Chronic Wasting Disease, and Creutzfeldt-Jakob disease (CJD). Prions (short for proteinaceous infectious particle) are known to have self-propagating or seeding activity. This means that misfolded prion proteins can cause healthy proteins to misfold and aggregate, which is thought to be a major method of pathogenesis in prion diseases. RT-QuIC is a very sensitive and specific test for CJD.
RT-QuIC works to detect CJD by taking a cerebrospinal fluid sample from a biopsy or necropsy and combining it with recombinant prion protein (rPrP) and thioflavin T. The specific type of rPrP used is recombinant truncated hamster PrP, but more on that later. In the presence of PrPSc, rPrP changes shape and forms fibrils which bind the thioflavin T. Bound thioflavin fluoresces, and the amount of fluorescence is measured over 30 hours. There is usually a significant lag phase of 5 to 20 hours before fluorescence is detectable, and this is what makes the assay take so long. If it were only performed for 10 or 20 hours, a significant number of cases would go undetected. Though this test is time-consuming, it boasts a sensitivity of 92% and a specificity of 100%. Sensitivity is the probability that a positive result corresponds with a true positive, and specificity is the probability that a negative result is a true negative.
Some concerns that can affect sensitivity and specificity are the type of rPrP used, white and red cell counts, and protein concentration. White cell counts above 10 x 10⁶ and red cell counts above 1250 x 10⁶ can interfere with interpretation, as can protein concentrations above 1 g per liter. The type of rPrP used is also a major factor. Recombinant prion protein can be of human or hamster origin, and it can be truncated or full-length. Truncated hamster rPrP yields similar sensitivity to other types while only taking 30 hours versus 90 hours for any other option. This makes it the most efficient and time-saving choice, and makes the method much “QuIC”-ker than it would be otherwise.
