Microplate ELISA for detection of antibodies to DNA in patients with systemic lupus erythematosus: specificity and correlation with Farr radioimmunoassay.
A 96-well microplate ELISA for the detection of antibodies to DNA is described. A number of buffers and precoating treatments were used to evaluate the optimal method for coating the plate with DNA. These included pretreatment of the plates with poly-L-lysine or protamine sulfate, and posttreatment with glutaraldehyde, none of which improved the performance of the assay. Whereas bicarbonate and borate coating buffers gave equivalent and satisfactory results, TRIS buffer resulted in very high binding of immunoglobulin to wells not coated with antigen. Sera from groups of patients with autoimmune disease as well as normal sera were tested against plates optimally coated with native E. coli DNA, calf thymus DNA, and heat-denatured DNA. Using native E. coli DNA, virtually none of 35 normal sera had any detectable antibody. With this antigen, as well as with native calf thymus DNA, significant levels of DNA antibody were found only in SLE patients. Most patients with SLE or drug-induced lupus, as well as some patients with rheumatoid arthritis and normal individuals had antibodies that bound to heat-denatured (single-stranded) DNA. Using either native E. coli or calf thymus DNA, a good correlation was found between the amount of DNA antibody detected by ELISA and the Farr-type radioimmunoassay.
Sutjita
M
M
Department of Clinical Immunology, Flinders Medical Centre, Bedford Park 5042, South Australia.
Hohmann
A
A
Boey
M L
ML
Bradley
J
J
D001323:Autoantibodies / Q000032:analysis*; D004247:DNA / Q000276:immunology*; D004277:DNA, Single-Stranded / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D008180:Lupus Erythematosus, Systemic / Q000276:immunology*; D011863:Radioimmunoassay / Q000379:methods
D001323:Autoantibodies; D004277:DNA, Single-Stranded; D004247:DNA