Submitted to: Hybridoma
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: July 16, 2012
Publication Date: October 25, 2012
Citation: Stanker, L.H., Scotcher, M.C., Lin, A.V., Mcgarvey, J.A., Prusiner, S., Hnasko, R.M. 2012. Novel epitopes identified by Anti-PrP monoclonal antibodies produced following immunization of Prnp0/0 Balb/cJ mice with purified scrapie prions. Hybridoma. 31(5):312-324. doi:10.1089/hyb.2012.0022. Interpretive Summary: Prion disorders represent a group of fatal neurological diseases caused by accumulation of the alternative folding of a normal cellular protein (PrP). Prion disease has been described in humans (Creutzfeldt-Jakob disease) and in animals (Bovine Spongiform Encephalopathy or BSE in cattle, and scrapie in sheep).Confirmation of prion disease relies on immunoassays and microscopic assessment of brain tissue. Immunoassays rely on the ability of antibodies to specifically react with an antigen or pathogen, in this case binding to the prion protein in the brain. Current tests, while able to detect abnormally folded protein, are not sensitive and rely on the digestion of the normal protein with enzymes. This research describes development of novel monoclonal antibodies that bind to specific regions of PrP. These antibodies can be incorporated into new prion diagnostic tests that should provide producers and regulators with improved tools to identify and better manage prion diseases such as BSE. They also will serve as highly specific tools to further study the biology of this unusual class of diseases.
Technical Abstract: Prions, or infectious proteins, cause a class of uniformly fatal neurodegenerative diseases. Prions are composed solely of an aberrantly folded isoform(PrPSc)of a normal cellular protein (PrPC). Shared sequence identity of PrPSc with PrPC has limited the detection sensitivity of immunochemical assays, as antibodies specific for the disease-causing PrPSc isoform have not been developed. Here we report the generation of three new monoclonal antibodies(MAbs)to PrP, which were isolated following immunization of Prnp0/0 Balb/cJ mice with highly purified PrPSc isolated from brain lipid rafts. Epitope mapping using synthetic PrP peptides revealed that the three MAbs bind different epitopes of PrP. The DRM1-31 MAb has a conformational epitope at the proposed binding site for the putative prion conversion co-factor “protein X.” The DRM1-60 MAb binds a single linear epitope localized to the beta–alpha loop region of PrP, whereas DRM2-118 binds an epitope that includes sequences within the octarepeat region and near the site of N-terminal truncation of PrPSc by proteinase K. Our novel anti-PrP MAbs with defined PrP epitopes may be useful in deciphering the conformational conversion of PrPC into PrPSc.