Figure 1.
The effect of substrate concentration on RT-QuIC.
RT-QuIC was performed using hamster rPrP substrate at concentrations of 20, 35, 45, and 60 µg/well. Reactions contained minimal salt (5.5 mM NaCl) and employed sCJD M/V brain homogenate (BH) diluted 10−4 to seed conversion.
Figure 2.
The effect of pH and salt on RT-QuIC when substrate concentration is 60 µg/well.
(A) RT-QuIC was performed using reaction mixtures at pH 5.8 and 7.0 with minimal salt (5.5 mM NaCl). (B) RT-QuIC was performed using various concentrations of NaCl in the reaction mixture. PBS in the reaction mixture contributed 138 mM NaCl, and resulted in final salt concentrations of 188, 238, 288, and 338 mM where the additional NaCl concentrations were 50, 100, 150, and 200 mM, respectively. Reactions employed hamster rPrP at a concentration of 60 µg/well as the substrate and sCJD M/V brain homogenate (BH) at indicated dilutions to seed conversion.
Figure 3.
The effect of batch variability and seed genotype on RT-QuIC.
RT-QuIC was performed using two different batches, Batch #1 (A) and Batch #2 (B), of hamster rPrP. Both assays employed hamster rPrP at a concentration of 60 µg/well as the substrate and sCJD M/V brain homogenate (BH) at the indicated dilutions as the seed. (C) RT-QuIC was performed using hamster rPrP Batch #1 at a concentration of 60 µg/well as the substrate and sCJD M/M brain homogenate (BH) at indicated dilutions to seed conversion. Reactions contained minimal salt (5.5 mM NaCl).