Study of anti-DNA antibodies prepared by DNA cellulose or Cibacron blue chromatography.
We prepared anti-DNA antibodies from sera of lupus patients by either DNA cellulose or by Cibacron blue chromatography. Eluates from both columns were studied with respect to recovery of IgG, recovery, purification and specificity of anti-DNA activity. An attempt was made to raise rabbit anti-idiotypic antibodies against both eluates. DNA cellulose chromatography--if DNA leakage was prevented--yielded 58% recovery and 58-fold purification of the anti-DNA activity present in the original purified IgG sample. 1% of loaded IgG was recovered. Cibacron blue chromatography yielded 32% recovery and 1.1-fold purification of the anti-DNA activity. 29% of loaded IgG was recovered. Eluates of Cibacron blue were not pure as shown by their high binding activity against an unrelated antigen, tetanus toxoid. Eluates from DNA cellulose were pure and did not show anti-tetanus toxoid activity. Rabbit anti-idiotypic antibodies could be raised only against eluates of DNA cellulose suggesting that the eluates of Cibacron blue did not contain enough idiotypes to induce anti-idiotypic antibodies. The characterization of the rabbit anti-idiotypic antibodies showed that it contained two populations, one against site-specific idiotypes and the other against framework idiotypes. Anti-DNA antibodies prepared by Cibacron blue had idiotypes similar to those prepared by DNA cellulose. The present study demonstrates that DNA cellulose chromatography--if leakage of DNA is prevented--can yield excellent recovery and purification of anti-DNA activity. Anti-DNA antibodies prepared by DNA cellulose were enriched and could induce anti-idiotypic antibodies in rabbits. Tube chromatography on Cibacron blue yielded poor recovery and minimal enrichment of anti-DNA activity.
D000818:Animals; D000974:Antibodies, Antinuclear / Q000302:isolation & purification*; D002482:Cellulose / Q000031:analogs & derivatives*; D002846:Chromatography, Affinity / Q000379:methods*; D004247:DNA / Q000031:analogs & derivatives* / Q000276:immunology*; D003851:Deoxyribonucleases / Q000494:pharmacology; D006801:Humans; D007074:Immunoglobulin G / Q000302:isolation & purification; D007130:Immunoglobulin Idiotypes / Q000276:immunology; D011817:Rabbits; D014227:Triazines
D000974:Antibodies, Antinuclear; C033190:DNA-cellulose; D007074:Immunoglobulin G; D007130:Immunoglobulin Idiotypes; D014227:Triazines; C012842:Cibacron Blue F 3GA; D002482:Cellulose; D004247:DNA; D003851:Deoxyribonucleases