The Intact PTH Immunoassay is a twosite ELISA [EnzymeLinked ImmunoSorbent Assay] for the measurement of the biologically intact 84 amino acid chain of PTH. Two different goat polyclonal antibodies to human PTH have been purified by affinity chromatography to be specific for well defined regions on the PTH molecule. One antibody is prepared to bind only the midregion and Cterminal PTH 3 984 and this antibody is biotinylated. The other antibody is prepared to bind only the Nterminal PTH 134 and this antibody is labeled with horseradish peroxidase [HRP] for detection. In this assay, calibrators, controls, or patient samples are simultaneously incubated with the enzyme labeled antibody and a biotin coupled antibody in a streptavidin coated microplate well. At the end of the assay incubation, the microwell is washed to remove unbound components and the enzyme bound to the solid phase is incubated with the substrate, tetramethylbenzidine (TMB). An acidic stopping solution is then added to stop the reaction and converts the color to yellow. The intensity of the yellow color is directly proportional to the concentration of intact PTH in the sample. A dose response curve of absorbance unit vs. concentration is generated using results obtained from the calibrators. Concentrations of intact PTH present in the controls and patient samples are determined directly from this curve.