In vitro amplification of misfolded prion protein using lysate of cultured cells

Charles E. Mays, Jihyun Yeom, Hae Eun Kang, Jifeng Bian, Vadim Khaychuk, Younghwan Kim, Jason C. Bartz, Glenn C. Telling, Chongsuk Ryou

Research output: Contribution to journalArticle

20 Scopus citations

Abstract

Protein misfolding cyclic amplification (PMCA) recapitulates the prion protein (PrP) conversion process under cell-free conditions. PMCA was initially established with brain material and then with further simplified constituents such as partially purified and recombinant PrP. However, availability of brain material from some species or brain material from animals with certain mutations or polymorphisms within the PrP gene is often limited. Moreover, preparation of native PrP from mammalian cells and tissues, as well as recombinant PrP from bacterial cells, involves time-consuming purification steps. To establish a convenient and versatile PMCA procedure unrestricted to the availability of substrate sources, we attempted to conduct PMCA with the lysate of cells that express cellular PrP (PrPC). PrPSc was efficiently amplified with lysate of rabbit kidney epithelial RK13 cells stably transfected with the mouse or Syrian hamster PrP gene. Furthermore, PMCA was also successful with lysate of other established cell lines of neuronal or non-neuronal origins. Together with the data showing that the abundance of PrPC in cell lysate was a critical factor to drive efficient PrPSc amplification, our results demonstrate that cell lysate in which PrPC is present abundantly serves as an excellent substrate source for PMCA.

Original languageEnglish (US)
Article numbere18047
JournalPloS one
Volume6
Issue number3
DOIs
StatePublished - 2011

All Science Journal Classification (ASJC) codes

  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)
  • General

Fingerprint Dive into the research topics of 'In vitro amplification of misfolded prion protein using lysate of cultured cells'. Together they form a unique fingerprint.

  • Cite this

    Mays, C. E., Yeom, J., Kang, H. E., Bian, J., Khaychuk, V., Kim, Y., Bartz, J. C., Telling, G. C., & Ryou, C. (2011). In vitro amplification of misfolded prion protein using lysate of cultured cells. PloS one, 6(3), [e18047]. https://doi.org/10.1371/journal.pone.0018047