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Development of method for the measurement of parathyroid hormone and its fragments based on immunocapture and mass spectrometry (IMUNO-MS)

Abstract

Abstract: parathormone (PTH) is a linear peptide with 84 amino acids (AA) produced exclusively by the parathyroid glands that have a fundamental role in calcium and phosphorous metabolism. Its biological activity is restricted to the first AA of the amino terminal portion of the molecule, a short half-life (2-3 min), and a complex intra and extra cellular metabolism. Several circulating forms, biologically active or inactive are present in circulation. Since the description of the first radioimmunoassays in the 1960's (1st generation assays), till the introduction in the 1990's of immunometric assays (2nd generation assays), assay specificity has been a constant problem. Not even the description of immunometric assays employing antibodies direct against the first AA (3rd generation assays), brought the expected improvement. In 1995 the technic of immune-MS was described, relying on an immunoadsorption, followed by mass spectrometric reading. In 2010 the first assays for PTH utilizing those principles were described (4th generation assays). In this project our objective is the development of an immuno-MS method based on a carboxyl terminal specific monoclonal antibody, followed by enzymatic (tryptic) digestion and mass spectrometric reading. Samples to be studied include patients with primary and secondary (to chronic Kidney Disease, CKD) hyperparathyroidism. In this late group we intend, with the discrimination and quantification of the circulating peptide forms, active or inactive, to optimize clinical and surgical conducts, particularly in patients with mineral and bone disorders (CKD-MBD). (AU)