A method for high yield isolation and purification of anti-native DNA antibodies present in lupus sera.
Antibodies to nucleic acids may serve as biochemical tools or as probes of cellular function. Particularly important, but also particularly difficult to obtain, is antibody which reacts exclusively with double stranded DNA. We describe here a method for the separation of antibodies to double stranded DNA from SLE serum, using hydroxyapatite to which DNA is adsorbed at a low molarity of phosphate buffer. Having applied the serum to the column we passed it through a continuous gradient of phosphate buffer ranging from 0.005 to 0.5 M. Deoxyribonuclease and magnesium ions were added when the gradient had reached the molarity at which single stranded DNA had already been desorbed and double stranded DNA began to be eluted. The antibody to native DNA that we obtained reacted in complement fixation, counterimmunoelectrophoresis and Farr's assay with native DNA and did not react with single stranded DNA, single and double stranded RNA or with a panel of 24 protein-coupled nucleosides, nucleotides and dinucleotides.
D000906:Antibodies / Q000302:isolation & purification*; D000915:Antibody Affinity; D000918:Antibody Specificity; D001666:Binding Sites, Antibody; D002845:Chromatography; D002850:Chromatography, Gel; D003168:Complement Fixation Tests; D004247:DNA / Q000276:immunology*; D006801:Humans; D006882:Hydroxyapatites; D007074:Immunoglobulin G; D008180:Lupus Erythematosus, Systemic / Q000276:immunology*
D000906:Antibodies; D006882:Hydroxyapatites; D007074:Immunoglobulin G; D004247:DNA