As hyperinsulinaemia has been shown to be a risk factor for non-insulin-dependent diabetes, cardiovascular disease and hypertension, the measurement of serum insulin levels may provide an additional early screening test for these diseases. Biological assaying of insulin is now facilitated by the IMx-Abbott kit, an enzyme immunoassay which does not cross-react with proinsulin and thus provides more specific insulin determination than conventional methods. Fasting insulin concentrations were determined in a population of 4,032 men and women 30 to 64 years of age, all volunteers for a medical check-up. Concentrations were slightly higher for men in all age-classes (median values of 5.9 and 5.4 microU/ml respectively for men and women). Although significant differences were found in serum insulin concentrations between the four age-classes for men and women, there were no significant differences between the three age-classes for men up to 59 years (median: 5.8 microU/ml, 95th percentile 14.0 microU/ml) or between the two age-classes for women up to 49 years (5.2, 12.5). Fasting concentrations were increased above these age thresholds: men (6.4, 15.6), women (5.6, 14.0). The reference population consisted of 3,081 non-diabetic, glycosuria-negative subjects with a body mass index and glucose concentration lower than the 95th percentiles for their age and sex. The reference values for fasting insulin concentrations were: 1) women 30-49 years: median 5.1 microU/ml (95% confidence interval: 4.9-5.3), 95th percentile 11.2 microU/ml (10.9-11.9); and 2) men 30-64 years and women 50-64 years: median 5.6 microU/ml (5.4-5.7), 95th percentile 12.6 microU/ml (12.0-13.0).