Principle of Assay
The Human Anti-Denosumab Antibody ELISA Kit (A326365) is based on a double-antigen sandwich ELISA principle for the qualitative detection of anti-denosumab antibodies in human samples. The assay utilizes the specific binding of anti-denosumab antibodies to denosumab antigen. Denosumab antigen is pre-coated onto a 96-well microplate. During sample incubation, anti-denosumab antibodies present in the sample bind to the immobilized antigen. A biotinylated denosumab antigen is subsequently added, allowing formation of a sandwich complex with the captured antibodies. After washing to remove unbound components, HRP-conjugated streptavidin is added and binds to the biotinylated antigen. Excess conjugate is removed by washing, and TMB substrate solution is then added, resulting in a blue color generated by the HRP enzymatic reaction. The reaction is terminated by the addition of an acidic stop solution, producing a yellow color. Absorbance is measured at 450 nm using a microplate reader. Results are interpreted qualitatively by comparing sample absorbance values to the assay controls to determine the presence of anti-denosumab antibodies.
