Enzyme-Linked Immunosorbent Assay Method for Human Autophosphorylated Insulin Receptor: Applicability to Insulin-Resistant States

Abstract

The insulin receptors from erythrocytes of 50 patients with non-insulin-dependent diabetes mellitus were tested for their ability to autophosphorylate. The assay was performed by a new enzyme-linked immunosorbent assay system that used monoclonal anti-insulin receptor antibodies absorbed to microtiter plates as a first antibody and polyclonal antiphosphotyrosine antibody as a labeled second antibody. By this assay, 3 patients were identified with defects in their insulin receptor kinase, although their defects appeared heterogeneous. Patient 1 had 85% less maximal autophosphorylation with a normal ED₅₀ (1.6 × 10⁻⁹ M insulin). Patient 2, who had polycystic ovary disease, had a 49.2% decrease in maximal autophosphorylation of insulin receptors, and the ED₅₀ was shifted to the right (5.6 × 10⁻⁸ M). Patient 3 with acanthosis nigricans had a normal maximal autophosphorylation, but the ED₅₀ shifted to the right (2.9 × 10⁻⁸ M). The mechanisms for the diversity detected in this assay is not known, but this technique has sufficient specificity and sensitivity to be used to screen for insulin-resistant patients who have a lack of kinase activity.