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Creutzfeld-Jakob disease(CJD)and Gerstmann-Straussler syndrome(GSS)of humans as well as scrapie of animals are caused by prions.The scrapie prion protein isoform(PrPSc)in the only macromolecule identified to date which is a component of the infectious prior particle.PrPSc is converted to PrP 27-30 by limited proteolysis while the cellular isoform,designated PrPC,is completely digested under the same conditions.ELISA studies demonstrated that native PrP 27-30 bound to plastic surfaces resisted proteolysis and exhibited little or no immunoreactivity but after denaturation with guanidinium thiocyanate(GdnSCN),immunoreactivity was greatly enhanced.PrPSc bound to nitrocellulose also exhibited enhanced immunoreactivity after denaturation.PrPSc was readily detected in brain extracts from scrapie-infected hamsters,mice,and sheep by dot-blot immunoassays using limited proteolysis followed by GdnSCN denaturation.The high sensitivity and specificity of the immunoassay allowed detection of regional differences in PrPSc in sheep brain.CJD prior protein isoform (PrPCJD)was also detected in the brains of all 10 patients tested with neuropathologically confirmed CJD and in 1 patient with GSS.Enhanced immunoreactivity of PrPSc or PrPCJD after denaturation cannot only be used for immunodiagnosis of prion diseases but may also form the basis of new assays in experimental studies directed at the chemical structure of the prion particle. |
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