Formalin RT-QuIC assay detects prion-seeding activity in formalin-fixed brain samples from sporadic Creutzfeldt-Jakob...

The neuropathology of sporadic Creutzfeldt-Jakob disease (sCJD) is usually investigated using formalin-fixed and formic acid-treated brain tissue. However, formalin and formic acid treatment can interfere with immunostaining of abnormal prion protein. Therefore, there is a need for biochemical methods other than immunostaining to investigate abnormal prion protein in postmortem tissue. We developed RT-QuIC to quantitate the seeding activity (SD₅₀) of sCJD brain tissue treated with formalin and formic acid.

We used endpoint RT-QuIC assays to analyze SD₅₀ in formalin-fixed brain tissue from 19 sCJD patients (14 MM1 cases, 3 MM2-thalamic form [MM2T] cases and 2 MM2-cortical form [MM2C] cases) diagnosed according to Parchi's classification. We assessed SD₅₀ in brains after incubation in formalin solution for over 1 month, and after treating formalin-fixed brain tissue with formic acid. We also examined how the SD₅₀ values from formalin-fixed brain samples compared with neuropathological and immunohistochemical findings.

The SD₅₀ values of formalin-fixed brain samples from 14 MM1 cases, 2 MM2C cases, and 2 MM2T cases were 10^7.77±0.57/g tissue, 10^7.44±0.24/g tissue and 10^6.00±0.77/g tissue, respectively. The average SD₅₀ value in MM1 unfixed brains decreased by 10^2.04 after formalin fixation for 1 month. In MM1 cases, after combined formalin and formic acid treatment, the SD₅₀ value was reduced by approximately 10^5.16 compared with that of unfixed tissue. The SD₅₀ values of formalin-fixed tissue showed a consistent pattern with the neuropathological findings in most brain regions examined.

RT-QuIC enables the study of formalin-fixed brain tissue from sCJD patients that has not previously been amenable to analysis.