Insulin assays play a central role in the investigation of glucose metabolism disorders (investigation of the causes of hypoglycemia, assessment of beta-cell function and determination of pathogenesis of type 1 and type 2 diabetes), and in studies on the pharmacology of insulin itself. Greater convenience and improved reproducibility, sensitivity and specificity have been achieved with human insulin immunometric assays. Besides human insulin, insulin analogues (lispro, aspart and glargine) have been introduced to therapeutic use. The specificity of human insulin assays to these analogues should be assessed. Anti-insulin antibodies are present in a significant proportion of sera to be analyzed for insulin. The sensitivity of insulin assays to interference from these antibodies should also be assessed. Interferences from anti-insulin antibodies and hemolysis, which degrades insulin molecules, remain the main pitfalls of insulin assays. Immunometric assays have also improved the sensitivity and reproducibility of free insulin measurements. Standardization of insulin immunoassays is still called for.