Adrenocorticotropic Hormone (ACTH)
The ACTH test measures plasma adrenocorticotropic hormone, the anterior pituitary peptide that drives adrenal cortisol synthesis. It is the key test for distinguishing ACTH-dependent from ACTH-independent Cushing syndrome and for separating primary from secondary adrenal insufficiency.
1
Biomarkers
Blood (plasma)
Sample
3–5 business days
Turnaround
No
Fasting
What is the ACTH test?
The Adrenocorticotropic Hormone (ACTH) test measures plasma concentrations of the pituitary peptide that regulates adrenal cortisol production. It is the decisive investigation for localising disorders of the hypothalamic–pituitary–adrenal axis, distinguishing whether a cortisol abnormality originates in the adrenal gland, the pituitary, or the hypothalamus. Practitioners order ACTH to classify hypercortisolism once biochemical Cushing syndrome has been confirmed, to determine the cause of adrenal insufficiency, and occasionally to investigate suspected ectopic ACTH secretion or congenital adrenal hyperplasia. The test is not a screening tool; it follows abnormal cortisol or synacthen-stimulation results and must be interpreted alongside a simultaneously drawn cortisol to be meaningful. Its extremely short plasma half-life and sensitivity to pre-analytic degradation make specimen handling as important as the assay itself.
What does the ACTH test measure?
Adrenocorticotropic hormone is a 39-amino-acid polypeptide synthesised and secreted by corticotroph cells of the anterior pituitary. It is cleaved from the larger precursor pro-opiomelanocortin and drives adrenal cortisol synthesis by binding to melanocortin-2 receptors on the zona fasciculata. ACTH secretion follows a pronounced circadian rhythm, peaking in the early morning hours and reaching a nadir in late evening. Its release is pulsatile, governed by hypothalamic corticotropin-releasing hormone and subject to negative feedback by circulating cortisol, forming the hypothalamic–pituitary–adrenal axis.
Plasma ACTH is the central test for distinguishing ACTH-dependent from ACTH-independent causes of hypercortisolism and for diagnosing primary and secondary adrenal insufficiency. An elevated ACTH with low cortisol points to primary adrenal failure; a suppressed or inappropriately normal ACTH with low cortisol suggests secondary or tertiary insufficiency. In Cushing syndrome, a detectable or elevated ACTH indicates a pituitary or ectopic source, while suppressed ACTH implicates an adrenal tumour. Because of its short half-life, lability, and circadian variation, ACTH measurement demands rigorous pre-analytic handling and careful interpretation in the context of paired cortisol.
Why is the ACTH test ordered?
- Classification of confirmed biochemical Cushing syndrome, to distinguish ACTH-dependent (pituitary or ectopic) from ACTH-independent (adrenal) hypercortisolism
- Diagnosis and differentiation of adrenal insufficiency, pairing ACTH with basal or stimulated cortisol to separate primary from secondary or tertiary causes
- Investigation of suspected ectopic ACTH secretion in patients with rapid-onset Cushing syndrome, hypokalaemic alkalosis, or known neuroendocrine tumours
- Evaluation of equivocal synacthen-test results, where the ACTH–cortisol pair clarifies the axis defect
- Monitoring of Nelson syndrome in patients who have undergone bilateral adrenalectomy for Cushing disease
- Assessment of congenital adrenal hyperplasia when 17-hydroxyprogesterone is elevated, to document the compensatory ACTH rise
- Work-up of incidental adrenal masses, where paired ACTH and cortisol help determine autonomous cortisol secretion
Sample collection and turnaround
Sample type
Blood (plasma)
Fasting required
No
Collection method
Venipuncture at patient service centre
Turnaround
3–5 business days
Collection notes
Phlebotomy into a chilled EDTA tube at the patient service centre, ideally between 0700 and 0900 hours to capture the morning peak. The patient should be seated and rested for 15–30 minutes before the draw to minimise stress-related elevation. Collect cortisol simultaneously in a separate serum or heparinised plasma tube. Centrifuge immediately in a refrigerated centrifuge, separate plasma, and freeze within 30 minutes; ACTH degrades rapidly at room temperature. Transport on dry ice if not analysed locally. Instruct patients to withhold morning glucocorticoid replacement (if applicable) until after the draw.
Specimen requirements
Minimum 1 mL EDTA plasma, chilled and frozen within 30 minutes. Note the exact time of collection. Paired cortisol specimen mandatory for interpretation. ACTH is unstable; pre-analytic mishandling is the most common cause of spuriously low results.
What can affect ACTH results?
ACTH follows a pronounced diurnal rhythm, highest at 0600–0800 hours and lowest in late evening; non-standardised timing makes interpretation unreliable, so early-morning sampling is essential for basal measurements.
ACTH is cleaved rapidly by plasma peptidases; delay in chilling, centrifugation, or freezing can lower measured values by 20–30 per cent within an hour. Collection into chilled EDTA tubes, immediate centrifugation in a refrigerated centrifuge, and prompt freezing are mandatory.
Physical or psychological stress activates the HPA axis and raises ACTH transiently; collecting a basal sample in an anxious or acutely unwell patient can yield spuriously elevated results.
Exogenous corticosteroids suppress ACTH through negative feedback; patients on replacement or pharmacological glucocorticoids must withhold the morning dose before the test to avoid suppression, and those on long-term therapy may have prolonged axis suppression.
Oestrogens raise corticosteroid-binding globulin, which can modestly elevate total cortisol and lead to compensatory ACTH changes; interpretation in pregnancy or on oestrogen should account for altered cortisol dynamics.
Immunoassays may cross-react with ACTH precursors or fragments or encounter interference from heterophile antibodies; discordant results warrant measurement by a second-generation or mass-spectrometry method.
What do ACTH results mean?
Elevated
Primary adrenal insufficiency (Addison disease, congenital adrenal hyperplasia, adrenalitis), where impaired cortisol synthesis removes negative feedback. Ectopic ACTH syndrome from neuroendocrine tumours (small-cell lung cancer, carcinoid, phaeochromocytoma). Cushing disease (pituitary adenoma). Nelson syndrome post-adrenalectomy. Rarely, familial glucocorticoid deficiency or ACTH receptor mutations.
Normal
A normal ACTH is meaningful only in the context of paired cortisol. An ACTH in the reference range with a high cortisol suggests ACTH-dependent hypercortisolism; the same ACTH with a low cortisol may indicate partial secondary insufficiency. Time of sampling and clinical context are essential.
Reduced
ACTH-independent Cushing syndrome due to adrenal adenoma or carcinoma, where autonomous cortisol suppresses the pituitary. Secondary (pituitary) or tertiary (hypothalamic) adrenal insufficiency from tumour, surgery, radiation, infiltration, or apoplexy, or from prolonged exogenous glucocorticoid therapy. Long-term suppression can persist for months after stopping glucocorticoids.
Note: ACTH must be collected under rigorous pre-analytic conditions — chilled EDTA plasma, immediate centrifugation, and freezing — or results will be spuriously low. It must be interpreted with a simultaneously drawn cortisol; isolated ACTH values are rarely diagnostic. Diurnal variation means that timing matters, and stress or glucocorticoid therapy can render a single sample uninterpretable. Dynamic tests (CRH stimulation, high-dose dexamethasone suppression) are often required to complete the differential diagnosis.
Frequently asked questions
ACTH drives cortisol synthesis and is regulated by cortisol feedback, so interpreting one without the other is meaningless. The paired values determine whether cortisol abnormalities originate in the adrenal or the pituitary.
How to access the ACTH test
The ACTH reference page is free and open to all practitioners. Create a free practitioner account to see pricing, save tests to your list, and generate requisition forms. Verification takes 1–2 business days.
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Published: October 7, 2026
Last reviewed: October 7, 2026
- Nieman LK, et al. The diagnosis of Cushing's syndrome: an Endocrine Society clinical practice guideline. J Clin Endocrinol Metab. (2008)
- Bornstein SR, et al. Diagnosis and treatment of primary adrenal insufficiency: an Endocrine Society clinical practice guideline. J Clin Endocrinol Metab. (2016)
- Raff H, Findling JW. A physiologic approach to diagnosis of the Cushing syndrome. Ann Intern Med. (2003)
- Kronenberg HM, et al. Williams Textbook of Endocrinology, 14th edition. Chapter on hypothalamic–pituitary–adrenal axis. (2020)
This content is for licensed healthcare practitioners only and does not constitute medical advice. Clinical decisions should be based on the full clinical picture and not on any single test result.