AJP - Endocrinology and Metabolism, Vol 257, Issue 3 E451-E457, Copyright © 1989 by American Physiological Society
Human insulin receptor radioimmunoassay: applicability to insulin-resistant states
V. Pezzino, V. Papa, V. Trischitta, A. Brunetti, P. A. Goodman, M. K. Treutelaar, J. A. Williams, B. A. Maddux, R. Vigneri and I. D. Goldfine
Cattedra di Endocrinologia, Universita di Catania, Ospedale Garibaldi, 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
greater than 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.
