Extraction and purification of active IgG from SSPE and MS brain.
Immunoglobulin (Ig) G was purified from soluble and membrane fractions of postmortem subacute sclerosing panencephalitis (SSPE) brain, multiple sclerosis (MS) brain plaque-periplaque white matter, and normal human brain (NHB) white matter. After homogenization in 0.32 M sucrose and removal of cell debris and nuclei by low-speed centrifugation, soluble and crude membrane fractions were separated by ultracentrifugation. After removal of sucrose by dialysis, IgG was isolated from the soluble fraction by protein A affinity chromatography. IgG was obtained from the membrane fraction by elution at low pH and purification from the eluate by protein A chromatography. Whereas very little IgG was in NHB white matter, significant levels of IgG were recovered from both SSPE and MS brain. Both immunocytochemical staining of measles virus-infected cells in tissue culture and protein immunoblotting of virus-infected cell lysates showed that the IgG from SSPE brain contained activity specific for measles virus protein. The abundance, purity and functional activity of IgG extracted from SSPE and MS brain indicate that IgG extracted from the brain of humans with an inflammatory disease of unknown etiology can be used to identify its corresponding antigen.
Owens
G P
GP
Department of Neurology, University of Colorado Health Sciences Center, Denver 80262, USA. gregory.owens@UCHSC.edu
Burgoon
M P
MP
Devlin
M E
ME
Gilden
D H
DH
D000818:Animals; D000914:Antibodies, Viral / Q000302:isolation & purification* / Q000378:metabolism; D001923:Brain Chemistry / Q000276:immunology*; D002460:Cell Line; D002462:Cell Membrane / Q000276:immunology; D002522:Chlorocebus aethiops; D003101:Collodion; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D015151:Immunoblotting; D007074:Immunoglobulin G / Q000302:isolation & purification* / Q000378:metabolism; D007525:Isoelectric Focusing; D007668:Kidney / Q000166:cytology; D008459:Measles virus / Q000276:immunology; D009103:Multiple Sclerosis / Q000276:immunology*; D019590:Nucleocapsid Proteins / Q000276:immunology; D011489:Protein Denaturation; D012995:Solubility; D013344:Subacute Sclerosing Panencephalitis / Q000276:immunology*
D000914:Antibodies, Viral; D007074:Immunoglobulin G; D019590:Nucleocapsid Proteins; D003101:Collodion