Two-Site Immunoradiometric Assay of Intact Salmon Calcitonin

Abstract

We developed a two-site immunoradiometric assay (IRMA) of salmon calcitonin (SCT) that detects intact SCT(1-32) and not peptide fragments of the hormone. This was accomplished by using monoclonal antibodies prepared against the peptide fragments SCT(1-11) and SCT(11-32). Two antibodies with specificity for each of the peptides were purified from their respective ascites and evaluated in a two-site format wherein one of the antibodies was adsorbed to polystyrene beads and the other was radioiodinated. In this assay format, the antibody pair detected intact SCT(1-32) but did not react with either SCT(1-11) or SCT(11-32). The sensitivity of the assay could be increased by exchanging the antibodies with respect to bead adsorption and radioiodination. Furthermore, by increasing the incubation time and volume of incubation of sample with the polystyrene-bead-adsorbed antibody, the effective detection limit of the assay could be improved. This assay system can be used to detect intact SCT when the presence of fragments of the hormone might otherwise complicate interpretation of assay data.