Monoclonal antibody for DNA measurement in biological fluids.
Two highly sensitive immunoassays for measuring soluble DNA are described. These methods employ a purified mouse monoclonal anti-DNA antibody in enzyme-linked immunoassays. One assay is an antigen capture method utilizing immobilized antibody. Bound DNA is subsequently detected with biotinylated monoclonal anti-DNA antibody, avidin-coupled horseradish peroxidase and a chromogenic substrate. The limit of detection of the assay was 2 ng/ml of DNA. A competition assay relying on immobilized heat-denatured DNA was also designed. In this assay the solution to be analyzed is mixed in equal proportions with monoclonal anti-DNA antibody and the mixture is incubated in the DNA-containing microtiter plates. The inhibition of antibody binding as detected with peroxidase-conjugated anti-mouse IgG was proportional to the concentration of DNA in the sample. The competition assay was tested for its ability to detect DNA in human plasma. The detection limit of DNA in plasma was approximately 150 ng/ml. The assays were compared in their ability to detect various size fragments of DNA. The competition assay was capable of detecting DNA fragments as small as 30 base pairs. In contrast, the capture assay failed to detect low molecular weight fragments up to 150 base pairs although its sensitivity for undigested DNA was comparable to the competition assay. The assay may be of use in the rapid quantitation of low levels of DNA, especially low molecular weight DNA and may also be useful in measuring DNA in human plasma.
D000911:Antibodies, Monoclonal; D001667:Binding, Competitive; D004247:DNA / Q000032:analysis* / Q000097:blood; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D006801:Humans; D008970:Molecular Weight; D009690:Nucleic Acid Conformation; D009707:Nucleosomes / Q000032:analysis
D000911:Antibodies, Monoclonal; D009707:Nucleosomes; D004247:DNA