A Two-Site Immunoradiometric Assay for Pth(1–84) Using N and C Terminal Specific Monoclonal Antibodies

Abstract
We report the development of a two-site immunoradiometric assay for PTH(1–84) based on two site-specific monoclonal antibodies—3B3 (radiolabelled antibody) specific for PTH(1–34) and ESQ1 (on solid phase) specific for PTH(74–84). Antibody 3B3 is sensitive to the oxidation of the methionine residues in PTH(1–34) therefore hydrogen peroxide (0·1 M) is added to the incubation mixture. Validation studies confirm quantitative recovery of both oxidized and reduced PTH(1–84). The assay has a minimum detection limit of 0·5 pmol/L and a range of 1·5–250 pmol/L with an intra-assay CV of less than 10% (2·8–250 pmol/L less than 5% CV). Studies on clinical samples indicate good discrimination between normal subjects (mean 2·21; range 1·0–5·0 pmol/L) and patients with primary hyperparathyroidism (mean 21·0; range 5·8–100 pmol/L) who in turn are well separated from patients with hypercalcaemia of malignancy (14/18 < 0·5 pmol/L).