Human insulin receptor radioimmunoassay: Applicability to insulin-resistant states(106 visite) Pezzino V, Papa V, Trischitta V, Brunetti A, Goodman PA, Treutelaar MK, Williams JA, Maddux BA, Vigneri R, Goldfine ID
Parole chiave: Insulin Receptor, Radioisotope, Somatomedin C Receptor, Controlled Study, Fibroblast, Human, Human Cell, Insulin Resistance, Leprechaunism, Lymphocyte, Methodology, Normal Human, Priority Journal, Radioreceptor Assay, Female, Placenta, Pregnancy, Radioimmunoassay, Support, Non-U.S. Gov, Non-U. S. Gov,
Cattedra di Scienza delle Costituzioni dell' Universita di Catania, Ospedale Garibaldi, Unita Sanitaria Locale 34, 95123 Catania, Italy
A radioimmunoassay of the human insulin receptor was developed employing a potent rabbit polyclonal antibody to the human insulin receptor and a highly purified human placental insulin receptor preparation. The receptor, obtained by sequential affinity chromatography with insulin receptor monoclonal antibody-agarose and wheat germ agglutinin-agarose, was radiolabeled with 125I-Bolton-Hunter reagent at specific activities of 2,100-3,300 Ci/mmol. Over 75% of this ligand was immunoprecipitable with the polyclonal antireceptor antibody and remained immunoprecipitable for > 45 days. The assay was sensitive to unlabeled receptor concentrations as low as 0.2 ng/0.5 ml; unlabeled insulin did not cross-react and unlabeled insulin-like growth factor (IGF)-I receptor cross-reacted weakly. The radioimmunoassay was applicable to the measurement of insulin receptors in tissues and cells that were extracted by solubilization in 1% Triton X-100; no purification of the extracted receptor was necessary. Of the three major target tissues for insulin action studied, liver had the highest concentration of receptors (47.6 ng/mg protein); fat and muscle had lower levels. Other studies with the radioimmunoassay indicated that insulin receptors were decreased both in monocytes from obese hyperinsulinemic subjects and in fibroblasts from patients with leprechaunism.
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