A method for the isolation of autoreactive B cells.
A technique was developed to isolate a population of autoreactive B cells from both normal and autoimmune-prone mice. Modifications of the procedure of Haas and Layton (1975) permitted coupling the nucleoside guanosine (GU) to gelatin and subsequently coating this matrix onto tissue culture dishes. After incubation on GU-gelatin, B lymphocytes specific for GU could be isolated. Specificity was demonstrated by rosetting techniques as well as by inhibition of binding to GU-gelatin by GU-containing conjugates. Isolated GU+ B cells were triggerable with GU-Brucella abortus antigen as well as LPS, to secrete anti-GU antibody in a direct plaque assay. The DNA-binding activity of the antibody was assessed using hapten inhibition of anti-GU PFC. Both native (N) DNA as well as denatured (D) DNA inhibited plaque formation. DNA-binding ability of secreted anti-GU antibody was also demonstrated by plaque formation using D-DNA-coated erythrocytes as target cells. Isolated GU+ B cells that are triggerable with antigen will be important in investigating growth, triggering and tolerance defects in a specific population of autoreactive B cells. In addition autoreactive B cells can now be compared to nonautoreactive hapten-specific lymphocytes. These properties as well as others can now be studied in controlled systems free from the regulatory effects of murine T or accessory cells.
D000818:Animals; D000921:Antibody-Producing Cells / Q000276:immunology; D000941:Antigens; D000955:Antigens, T-Independent / Q000276:immunology; D001324:Autoantigens; D001402:B-Lymphocytes / Q000166:cytology / Q000276:immunology*; D002469:Cell Separation / Q000379:methods; D004247:DNA / Q000276:immunology; D006151:Guanosine / Q000276:immunology*; D008180:Lupus Erythematosus, Systemic / Q000276:immunology*; D051379:Mice; D051381:Rats; D011947:Receptors, Antigen, B-Cell / Q000276:immunology
D000941:Antigens; D000955:Antigens, T-Independent; D001324:Autoantigens; D011947:Receptors, Antigen, B-Cell; D006151:Guanosine; D004247:DNA