Generation of monoclonal antibodies against prion proteins with an unconventional nucleic acid-based immunization strategy

Prion diseases belong to a group of neurodegenerative disorders affecting h umans and animals. The human diseases include kuru, Creutzfeldt-Jakob disea se (CJD), Gerstmann-Straussler-Scheinker syndrome (GSS) and fatal familial insomnia (FFI). The pathomechanisms of the prion diseases are not yet under stood. Therefore, monoclonal antibodies (mAbs) would provide valuable tools in diagnostics as well as in basic research of these diseases. In contrast to conventional strategies we have developed an immunization protocol base d on nucleic acid injection into non tolerant PrP0/0-mice. DNA or RNA codin g for different human prion proteins including the mutated sequences associ ated with CJD, GSS and FFP were injected into muscle tissue. The mice were primarily inoculated with DNA-plasmids encoding PRNP and boosted either wit h DNA, RNA or recombinant Semliki Forest virus (SFV) particles expressing P RNP. After hybridoma preparation, different mAbs against prion proteins wer e obtained and their binding behaviour was analysed by peptide-ELISA, Weste rn blot, immunofluorescence and immunoprecipitation. Our mAbs are directed against four different linear epitopes and may also recognize discontinuous regions of the native prion protein. It could, therefore, be demonstrated that immunization of non tolerant mice with DNA and live attenuated SF viru s is a valuable means to induce a broad immune response leading eventually to the generation of a panel of mAbs for basic science as well as for diagn ostics. (C) 1999 Elsevier Science B.V. All rights reserved.