Table 1.
Comparison of the 50-percent seeding doses (SD50) at 42°C.
Table 2.
Comparison of SD50 at 55°C.
Fig 1.
Amplification reactions with scrapie-infected hamster brain homogenates.
Results in Fig 1 are average fluorescent signals ± standard errors of the mean (SEM) for reactions at 42°C. The maximum fluorescence intensity was 260,000 relative fluorescent units (rfu) for all experiments. Fig 1A. RT-QuIC was conducted with scrapie-infected hamster brain homogenate serially diluted from 10−4 to 10−11 (10% w/v = 10−1). Fluorescence values for each dilution (4 wells/dilution) were measured with a FLUOstar Omega fluorimeter. Fluorescence readings were reported at different times as indicated. We reported fluorescent signals for control samples at the indicated time points. Fig 1B. QuIC on a Thermomixer-R conducted as in Fig 1A. The same microplate was read on an external fluorimeter at the indicated times and the average signal at each dilution was reported with its SEM. NBH = Ha-recPrP substrate seeded with normal hamster brain diluted 10−4. recPrP = Hamster recombinant PrP unseeded.
Fig 2.
Amplification reactions at 42°C.
Comparison of amplifications at 42°C with five TSE-infected brain suspensions as seeds. We used Ha-recPrP substrate to amplify PrPTSE from scrapie (Fig 2A), CWD (Fig 2B), macaque-adapted vCJD (Fig 2C) and sCJD (Fig 2D) brain suspensions. BV-recPrP substrate amplified human vCJD brain seed (Fig 2E). RT-QuIC fluorescence signals were recorded only at 18 hours and 60 hours (48 hours for sCJD). QuIC-R and QuIC-C signals were recorded at 18 hours and 24 hours, respectively. NBH = recombinant PrP (hamster or bank vole) seeded with normal brain diluted 10−4.
Fig 3.
Amplification reactions at 55°C.
Comparison of amplifications at 55°C with CWD and sCJD-infected brain suspensions as seeds at various reaction times. Fig 3A represents the average fluorescent signals and SEM with sCJD brain homogenates amplified with QuIC-C on Thermomixer-C at 8 hours and RT-QuIC on FLUOstar Omega at 8 hours and 24 hours. Fig 3B represents the average fluorescent signals and SEM with CWD brain homogenates amplified with QuIC-C on Thermomixer-C at 8 hours and 24 hours and RT-QuIC on FLUOstar Omega at 8 hours and 36 hours. NBH = hamster recombinant PrP seeded with normal brain diluted 10−4.