Free covalent aggregates of therapeutic insulin in blood of insulin-dependent diabetics.

Immunoreactive insulin (IRI) in the circulation of diabetics using insulin includes a 12,000-Mr covalent aggregate of insulin. In this study, both free and bound IRI were measured in nine type I diabetics who were treated sequentially for 3 wk with constant doses of conventional beef-pork, biosynthetic human, and beef-pork insulins (phases 1, 2, and 3, respectively). The aggregate accounted for 19 +/- 5.7, 38 +/- 7.7, and 26 +/- 7% of bound IRI, and 19 +/- 8.3, 35 +/- 7.3, and 31 +/- 5.0% of free IRI during phases 1, 2, and 3, respectively. Taken as concentrations (microU/ml), the absolute amounts of aggregate in the bound fraction were significantly (P less than .001) greater than those that were free, whereas the ratios of aggregate to monomer within each pool were similar (P greater than .1). The relative and absolute amounts of the insulin aggregate during each of the three treatment phases (compared by analysis of variance) were indistinguishable (P greater than 0.5). However, the aggregate was overestimated by a factor of approximately 2 when measurements were made on the basis of an insulin-monomer standard. We conclude that both the bound and free fractions of IRI contain insulin aggregate. Overreactivity of the aggregate in the radioimmunoassay contributes to the so-called hyperinsulinism of type I diabetes. As with insulin monomer, most of the 12,000-Mr aggregate is bound to antibodies. Because some of the aggregate is free, its biologic consequences must be assessed.