Blood prion seed amplification assays (RT-QuIC and PMCA in blood)

Key facts

• A 2016 PMCA study from the Mitchell Center in Houston, with the Carlo Besta institute in Milan and the Edinburgh surveillance unit, tested blood from 14 variant CJD cases and 153 controls, including sporadic CJD and other neurological patients, and reported 100 percent sensitivity and specificity, from a few microlitres of sample and in any blood fraction tested, with an estimated PrPSc concentration near 0.5 pg per ml.
• The same year a French team at the Etablissement Francais du Sang, with the Paris Brain Institute, the AP-HP CJD unit and the Edinburgh unit, reported a plasma amplification test for variant CJD with 100 percent sensitivity across 256 plasma samples and detection in two blood donors 1.3 and 2.6 years before symptoms began.
• A 2019 PMCA study in three macaques infected with macaque adapted variant CJD prions found the first blood sample positive 65 days after inoculation, which was 759 days before onset in two animals and 644 days in the third; per replicate sensitivity on 140 buffy coat samples was 95.8 percent and specificity 100 percent, with all 68 control replicates negative. Buffy coat worked better than plasma at the preclinical stage.
• A November 2023 Roslin study built whole blood IOME RT-QuIC, which captures prions from 200 microlitres of unfractionated whole blood on iron oxide magnetic beads before seeding a reaction with truncated ovine recombinant PrP. Nine of 12 blood samples from BSE infected sheep were positive, including samples taken about 24 months before clinical onset, and the work was funded by the UK Department of Health and Social Care through the NIHR.
• In the same study, bead PMCA was more sensitive than RT-QuIC on both matrices: on a reference human variant CJD brain homogenate RT-QuIC reached about a 10 to the minus 5 or minus 6 dilution against 10 to the minus 9 for bead PMCA, and in sheep blood bead PMCA called 11 positives against 8 by RT-QuIC. No human blood was tested.
• A blinded October 2024 comparison in Blood, led from Roslin with the French blood service, the MHRA, the Animal and Plant Health Agency and INRAE Toulouse, ran three assays on longitudinal sheep blood series: both PMCA versions reached 100 percent diagnostic sensitivity and RT-QuIC 61 percent, all three detected infection 26 months before clinical signs, and there were no false positives. Blood infectivity titres in a sensitive transgenic mouse line correlated with PrPSc detection.
• That 2024 work sits under BBSRC institute programme BBS/E/RL/230002C, which runs from March 2023 to March 2028 with 3,398,478 GBP, and the group has since taken 1.25 million euro of European Partnership on Animal Health and Welfare funding to adapt PMCA and RT-QuIC to tell prion strains apart.
• For sporadic prion disease, a March 2024 Japanese study using an enhanced QuIC with three human monoclonal antibodies, about 1000 fold more sensitive than standard RT-QuIC on brain tissue, found no seeding activity in whole blood, serum or plasma. The abstract says six patients, the methods and tables describe three, all MM1, and the paper does not explain the difference.
• Those authors ran roughly 1000 eQuIC reactions on blood and offer three explanations, that PrPSc in sporadic patients' blood is too scarce for current methods, that it may not be detectable at all, or that the antibodies do not suit blood; they favour the first two and point to tear fluid as an alternative route.
• Infectivity, as distinct from amplifiable seeds, has been found in human blood by bioassay: a 2014 brief report found infectivity in erythrocytes, leukocytes and plasma of one variant CJD patient and in the plasma of two of four sporadic CJD patients, at levels comparable to animals with transmissible spongiform encephalopathies.
• The UK route for judging such tests runs through the NIBSC CJD Resource Centre at South Mimms, whose stated purpose includes supplying reference material for blood screening tests, and whose staff are among the MHRA coauthors on the 2024 blinded comparison.
• The test that reads blood is not the test that reads spinal fluid: CSF RT-QuIC is an established diagnostic, while every blood result above is research, and nothing opened in this pass shows a blood assay offered as a clinical service.

Timeline

1. 2014Brief report finds infectivity by bioassay in blood of one variant CJD patient and in plasma of two of four sporadic CJD patients.
2. 2016Houston PMCA study reports 100 percent sensitivity and specificity on blood from 14 variant CJD cases; the French plasma amplification test is published in the same month and journal.
3. 2019Macaque PMCA study published, with blood positive 65 days after inoculation and more than 600 days before onset.
4. 2023Roslin whole blood IOME RT-QuIC paper published, detecting BSE in sheep blood about 24 months before onset.
5. 2023Roslin news item describes the bead based blood test and names the UK Department of Health and Social Care as funder.
6. 2024Japanese eQuIC study reports no detectable seeding activity in blood from sporadic prion disease patients.
7. 2024Blinded three assay comparison published in Blood, PMCA at 100 percent and RT-QuIC at 61 percent sensitivity, all three positive 26 months before signs.
8. 2026Roslin announces a 1.25 million euro programme to adapt PMCA and RT-QuIC for prion strain discrimination.