Estimating prion concentration in fluids and tissues by quantitative PMCA

Article metrics

Abstract

Prions, the proteinaceous infectious agent responsible for prion diseases, can be detected with high sensitivity by protein misfolding cyclic amplification (PMCA) technology. Here we describe a quantitative PMCA procedure to calculate the concentration of very low levels of prions in biological samples. Using this procedure, we determined the quantities of misfolded prion protein (PrPSc) in brain, spleen, blood and urine of scrapie-affected hamsters.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Figure 1: Relationship between PrPSc concentration and PMCA rounds required for detection.
Figure 2: Detection of PrPSc in spleen of scrapie-affected hamsters.

References

  1. 1

    Aguzzi, A. & Calella, A.M. Physiol. Rev. 89, 1105–1152 (2009).

  2. 2

    Soto, C. Nat. Rev. Microbiol. 2, 809–819 (2004).

  3. 3

    Brown, P. Neurology 70, 713–722 (2008).

  4. 4

    Wadsworth, J.D. et al. Lancet 358, 171–180 (2001).

  5. 5

    Saborio, G.P., Permanne, B. & Soto, C. Nature 411, 810–813 (2001).

  6. 6

    Saa, P., Castilla, J. & Soto, C. J. Biol. Chem. 281, 35245–35252 (2006).

  7. 7

    Castilla, J., Saa, P. & Soto, C. Nat. Med. 11, 982–985 (2005).

  8. 8

    Saa, P., Castilla, J. & Soto, C. Science 313, 92–94 (2006).

  9. 9

    Thorne, L. & Terry, L.A. J. Gen. Virol. 89, 3177–3184 (2008).

  10. 10

    Murayama, Y. et al. J. Gen. Virol. 88, 2890–2898 (2007).

  11. 11

    Gonzalez-Romero, D., Barria, M.A., Leon, P., Morales, R. & Soto, C. FEBS Lett. 582, 3161–3166 (2008).

  12. 12

    Morales, R. et al. Biochem. Biophys. Res. Commun. 377, 373–378 (2008).

  13. 13

    Castilla, J. et al. Methods Enzymol. 412, 3–21 (2006).

  14. 14

    Castilla, J., Saá, P., Hetz, C. & Soto, C. Cell 121, 195–206 (2005).

  15. 15

    Saa, P., Castilla, J. & Soto, C. Methods Mol. Biol. 299, 53–65 (2005).

Download references

Acknowledgements

We thank D. Gonzalez-Romero (University of Texas Houston Medical School) for participating in the PMCA experiments with urine, R. Diaz-Espinoza (University of Texas Houston Medical School) for providing recombinant PrP and D. Zhao (China Agricultural University) for suggestions and providing support to B.C. This research was supported in part by US National Institutes of Health grants R01NS049173 and P01AI077774 to C.S.

Author information

B.C. performed most of the experiments, analyzed the data and prepared the figures; R.M. helped with experimental design and data analysis; M.A.B. performed the PMCA experiments in blood; and C.S. developed the hypothesis, designed and analyzed all data, wrote the manuscript and supervised the project.

Correspondence to Claudio Soto.

Ethics declarations

Competing interests

C.S. is inventor of the PMCA technology and is founder, vice-president and chief scientific officer of Amprion, Inc, a biotechnology company focused on developing technologies to detect misfolded proteins associated with neurodegenerative diseases.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Chen, B., Morales, R., Barria, M. et al. Estimating prion concentration in fluids and tissues by quantitative PMCA. Nat Methods 7, 519–520 (2010) doi:10.1038/nmeth.1465

Download citation

Further reading