Monoclonal antibodies specific for the free alpha subunit of glycoprotein hormones and their use in the development of a sensitive immunofluorometric assay

Glicoprotein hormone free alpha subunit has been used as a marker for some pituitary tumors and to study the reactivity of glycoprotein hormone-producing cells under different circunstances. We describe a highly sensitive ans specific immunofluorometric assau for the measurement of serum free alpha subunit levels. The assay is based on a monoclonal antibody, specific for free alpha subunit, bound to microtiter plates. As tracer antibody we employed an europium-labelled free/complexed alpha subunit specific monoclonal antibody. Using overnight incubation and 50µl samples, the least detectable dose was of the order of 4 ng/1. Cross-reactivity with LH, TSH, FSH, and hCG was 6.5, 1.2, 4.3 and 1.1 percent, repectively. Normal adult males showed values ranging from 120 to 790ng/l, not different from normal adult premenopausal females (88 to 604 ng/l). In post-menopausal females, serum concentrations were significantly highler, ranging from 341 to 407 ng/l. In 56 patients with untreated pituitary tumors (18 "non-secreting", 25 GH-producing and 13 prolactin-producing tumors), 10 showed high values, 3 of them from the first group, 3 from the second and 4 from the third. We conclude that this highly sensitive assay can be a valualbe tool for the diagnosis and follow-up of selected patients with pituatary tumors and in other circumstances in which the glycoprotein hormone-producing cells of the pituitary require evaluation

Clinical utility of a 22-kDa growth hormone-specific assay

Human growth hormone (hGH) circulates in different molecular forms, with the 22-kDa monomer being the predominant one and the 20-k-Da variant corresponding to 5 to 15% of the serum hGH on a weight basis. Using monoclonal antibodies with different specificities we developed two immunoenzymometric assays, one with 22 + 20 k-Da specificity and the other specific only for the 22-kDa form. Both assays used microtiter plates as solid phase and streptavidin-peroxidase for color development; intra-assay CV was less than 10% in the range of 1 to 100 mlU/l for the 22 + 20 kDa assay and in the range of 3 to 100 for the 22-kDa assay, with an inter-assay CV of less than 14% for both assays, sensitivity was 0.2 mlU/l for the 22 + 20 kDa assay and 0.5 mlU/l for the 22-kDa assay. The two assays were compared by measuring 200 serum samples with detectable hGH levels by both assays. Higher values were obtained with the 22 + 20 kDa assay (62.1 ñ 59.2 ñ 6.1 mlU/l, mean ñ SD) with a correlation coefficient (r) of 0.99. In no clinical condition (28 patients with growth retardation and 14 acromegalics) did the two assays give discrepant values. We conclude that there was no practical advantage in using an assay with specificity restricted to the 22-kDa form for measuring hGH in clinical serum samples