Guanidine hydrochloride extraction and detection of prion proteins in mouse and hamster prion diseases by ELISA.
Current detection of transmissible spongiform encephalopathy (TSE) relies on the proteolytic generation of a protease-resistant core from the scrapie isoform of prion protein (PrP(Sc)) followed by immunoblotting. This process is non-quantitative, time-consuming, and technically demanding. Recently, an alternative in vitro test for TSE based on the differential extraction of brain homogenates using guanidine hydrochloride followed by DELFIA (Dissociation Enhanced Lanthanide FluoroImmunoAssay) has been developed. In the present study, this approach was adopted using a panel of anti-PrP monoclonal antibodies (MAbs) in conventional sandwich enzyme-linked immunosorbent assay (ELISA) to investigate hamster and two distinct strains of mouse prion diseases. Although PrP species were present in both soluble and insoluble fractions from normal as well as TSE samples, only the PrP species in the insoluble fractions from the latter samples were protease-resistant. In addition, certain anti-PrP MAb pairs could distinguish the PrP species in infected brains from those in the normal samples. The ability to differentiate disease-associated PrP isoforms without proteinase K digestion could serve as a panacea for developing a reliable and rapid diagnostic test for prion diseases.
Kang
Shin-Chung
SC
Institute of Pathology, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106, USA.
Li
Ruliang
R
Wang
Chuanping
C
Pan
Tao
T
Liu
Tong
T
Rubenstein
Richard
R
Barnard
Geoff
G
Wong
Boon-Seng
BS
Sy
Man-Sun
MS
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D001923:Brain Chemistry; D006224:Cricetinae; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D019791:Guanidine; D008647:Mesocricetus; D051379:Mice; D008815:Mice, Inbred Strains; D018620:PrPSc Proteins / Q000032:analysis / Q000276:immunology / Q000302:isolation & purification*; D017096:Prion Diseases / Q000175:diagnosis*; D011328:Prions / Q000032:analysis / Q000276:immunology / Q000302:isolation & purification; D012995:Solubility; D013048:Specimen Handling / Q000379:methods
D000911:Antibodies, Monoclonal; D018620:PrPSc Proteins; D011328:Prions; D019791:Guanidine