During recent studies with detergent-resistant membrane (DRM) preparations, we found that our published titers of Syrian hamster prions were incorrect (Figure 5B). From a re-examination of the data, we believe that some of the hamsters used in bioassays were diagnosed with scrapie before progressive CNS dysfunction occurred. When we used the time of death (Figure 5A) to calculate the prion titers, a more plausible set of data emerged. To do this, we used the equation:
Figure 5B.
Comparison of ID50 and specific infectivity in ill hamsters following intracerebral inoculation of PrPSc Brain as a 1% crude brain homogenate (25 µg), PrPSc DRM (1.35 µg) and purified PrPSc DRM-PK-PTA-G100 (0.8 µg). Isolation of PrPSc in DRMs from lipid rafts resulted in a >20-fold increase in specific infectivity relative to crude brain, but much less infectivity was found in the purified PrPSc DRM-PK-PTA-G100 fraction.
Log Titer = 25.33 − [12.47 × log(Z-61)] − logD
Where Z is the survival time in days, D is dilution factor and Titer is expressed in ID50 units per mL of the sample (1).
The results are summarized in new Figure 5B (below). As shown, the titer of the fraction denoted “PrPSc DRM-PK-PTA-G100” was 1.2 × 106 ID50 units/mL. While this fraction proved to be the most useful in raising anti-PrP antibodies, it exhibited much lower scrapie prion specific infectivity than we initially surmised.
We also found an error in Materials and Methods (page 100) describing the breeding of Prnp0/0/Balb/cJ mice; they were produced by backcrossing Balb/cJ mice with FVB/Prnp0/0 mice, not 129/SvJ/C57-Bl6 Prnp0/0 mice as stated.
References
- 1.Prusiner SB, Cochran P, Groth DF, Downey DE, Bowman KA, Martinez HM. Measurement of the scrapie agent using an incubation time interval assay. Ann Neurol. 1982;11:353–358. doi: 10.1002/ana.410110406. [DOI] [PubMed] [Google Scholar]