Immunology of DNA. I. The influence of reaction conditions on the Farr assay as used for the detection of anti-ds DNA.
The sensitivity and the specificity of the Farr assay for the detection of antibodies to double stranded (ds) DNA depends very much on the reaction conditions. The interaction between ds DNA and anti-ds DNA is inhibited when ionic strength and pH are increased. ds DNA is bound by normal sera at ionic strength lower than 0.11 M NaCl and at physiological ionic strength when the pH is lower than 7.2. Substantial binding of DNA by normal serum takes place in barbitone, borate or Tris-HCl buffers at concentrations of 30 mM or higher, even at a pH higher than 7.2. Such binding is due to Clq and is only partially prevented by heating the serum for 30 min at 56 degrees C, but 10 mM phosphate in the incubation mixture completely prevents it. Standardization of ionic strength, pH, phosphate concentration, incubation volume and DNA-serum ratio enhances the diagnostic usefulness of the Farr assay.
Immunology of DNA. I. The influence of reaction conditions on the Farr assay as used for the detection of anti-ds DNA. The sensitivity and the specificity of the Farr assay for the detection of antibodies to double stranded (ds) DNA depends very much on the reaction conditions. The interaction between ds DNA and anti-ds DNA is inhibited when ionic strength and pH are increased. ds DNA is bound by normal sera at ionic strength lower than 0.11 M NaCl and at physiological ionic strength when the pH is lower than 7.2. Substantial binding of DNA by normal serum takes place in barbitone, borate or Tris-HCl buffers at concentrations of 30 mM or higher, even at a pH higher than 7.2. Such binding is due to Clq and is only partially prevented by heating the serum for 30 min at 56 degrees C, but 10 mM phosphate in the incubation mixture completely prevents it. Standardization of ionic strength, pH, phosphate concentration, incubation volume and DNA-serum ratio enhances the diagnostic usefulness of the Farr assay.