The present study describes a radioimmunoassay for plasma adrenocorticotrophin (ACTH) in man with a sensitivity of at least 15 pg/ml.In-vitro studies using various fragments of ACTH showed immunobiological dissociation. However, this phenomenon does not appear to be of major importance in vivo, since stimulation and suppression tests showed overall parallel changes of immunoreactive plasma ACTH and plasma cortisol. The results obtained from patients with various forms of Cushing's syndrome are presented. It is concluded that basal plasma ACTH determinations are useful in the differential diagnosis of Cushing's syndrome and may help in predicting the development of a pituitary tumour after adrenalectomy.The supranormal plasma cortisol response to i.m. injection of lysinevasopressin in subjects with pituitary-dependent Cushing's syndrome is attributed both to an increased ACTH release by the pituitary and to an excessive response of the hyperplastic adrenal cortex. The i.v. infusion of dexamethasone at a rate of 1 mg/h for 5 h in subjects with pituitarydependent Cushing's syndrome was followed by significant and parallel decreases of both immunoreactive plasma ACTH and plasma cortisol. No rise of plasma ACTH levels was found with insulin-induced hypoglycaemia in subjects with untreated pituitary-dependent Cushing's syndrome.