Sensitivity for prion detection of SCEPA and mouse bioassay.

*<p>The ratio between PrP^Sc-positive and total wells is shown for one representative out of eight independent experiments.</p>†<p>Infectious titers were calculated with the Spearman-Karber formula <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1002538#ppat.1002538-Dougherty1" target="_blank">[55]</a> and expressed as tissue culture infectious units (TCIU)/g brain for SCEPA and LD₅₀ units/g brain for mouse bioassay, respectively.</p>$<p>Infectious titers were calculated using a GLM with binomial family complementary log-log link and expressed as mean log TCIU/g brain ± SE of 8 independent experiments for SCEPA and mean log LD50 units/g ± SE for two independent bioassays.</p>‡<p>Infectious titers were estimated for the combined two bioassays using a GLM regression with complementary log-log link function and expressed as log LD50 units/g brain.</p><p>The sensitivity for prion detection of SCEPA and mouse bioassay was determined by endpoint titration using RML mouse brain homogenate I6200. Aliquots of I6200 (10% (w/v), 9.3 log LD₅₀ units/g brain <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1002538#ppat.1002538-Cronier1" target="_blank">[7]</a>) were serially diluted 1∶10 into uninfected CD1 brain homogenate (10% w/v) in a range between 10⁻⁴ and 10⁻¹⁰. For mouse bioassay, groups of six Tga20 mice were inoculated intracerebrally with 30 µl of 1% (w/v) RML homogenates and attack rates and scrapie incubation times (Inc. time) were determined. In parallel experiments brain homogenates were diluted 1∶1000 into OFCS and cell layers of highly prion susceptible N₂aPK1-2 cells were infected with 300 µl aliquots. The input of prion infectivity for bioassay and SCEPA is expressed as mouse ic LD₅₀ units. A 10⁻⁷ dilution of I6200 corresponds to 200 LD₅₀ units/ml or 6 LD₅₀ units per 30 µl inoculum for the mouse bioassay and 60 LD₅₀ units per 300 µl per well for SCEPA, respectively. Infectious titers for SCEPA, expressed as TCIU/g brain represent mean values ± SE of 8 independent experiments. For mouse bioassay, two independent experiments are shown and titers are expressed as LD₅₀/g brain ± SE.