High-Dose ACTH Stimulation Test: Performance and Interpretation
The standard high-dose (250 µg) cosyntropin stimulation test should be performed by administering 250 µg cosyntropin IV or IM, measuring cortisol at baseline and 30 and 60 minutes post-injection, with a peak cortisol ≥500 nmol/L (18 µg/dL) excluding adrenal insufficiency, though newer specific assays may require lower cutoffs of 14-15 µg/dL. 1, 2
Test Protocol
Pre-Test Preparation
- Stop glucocorticoids and spironolactone on the day of testing (long-acting glucocorticoids may require longer washout periods) 2
- Discontinue estrogen-containing medications 4-6 weeks before testing to allow cortisol-binding globulin (CBG) levels to normalize, as elevated CBG falsely increases total cortisol measurements 2, 1
- The test can be performed at any time of day without time constraints, as results are not significantly affected by diurnal variation 1, 3
Dosing
Adults: 250 µg (0.25 mg) cosyntropin IV or IM 2
Pediatric patients: 2
- Birth to <2 years: 125 µg (0.125 mg)
- 2-17 years: 250 µg (0.25 mg)
Reconstitution
Reconstitute lyophilized powder with 1 mL of 0.9% sodium chloride to achieve final concentration of 0.25 mg/mL. Solution must be clear, colorless, and free of particulates. Discard if not used immediately. 2
Sample Collection
- Obtain baseline serum cortisol before injection
- Collect cortisol samples at exactly 30 and 60 minutes post-administration 2, 1
Interpretation
Standard Cutoff Values
Peak stimulated cortisol <500 nmol/L (18 µg/dL) at either 30 or 60 minutes is suggestive of adrenal insufficiency 1, 2. However, this threshold is based on older polyclonal immunoassays.
Assay-Specific Considerations
Critical caveat: Newer specific monoclonal antibody immunoassays (Roche Elecsys II) and LC-MS/MS measure cortisol concentrations approximately 21% lower than older assays 4, 5. For these newer assays, use cutoffs of 14-15 µg/dL (approximately 400-426 nmol/L) instead of 18 µg/dL to avoid false-positive diagnoses 5, 4.
The Endocrine Society guidelines note sensitivity of 92% (95% CI: 81-97%) for the high-dose test in primary adrenal insufficiency 1.
Timing of Measurement
The 60-minute cortisol value is superior to the 30-minute value alone. Using only a 30-minute measurement results in >13% false-positive rate, as many patients fail to reach the cutoff at 30 minutes but exceed it at 60 minutes 3, 6. A single 60-minute value ≥500 nmol/L is 99.7% concordant with traditional testing and can safely exclude adrenal insufficiency in >99% of cases 6, 3.
Supporting Diagnostic Information
When baseline cortisol is **<140 nmol/L (5 µg/dL)** combined with plasma ACTH elevated >2-fold above the upper reference limit (or >66 pmol/L), primary adrenal insufficiency is highly likely even before stimulation testing 1. Baseline cortisol <2 µg/dL predicts subnormal stimulated values 5.
Common Pitfalls and Conditions Affecting Results
Medications That Falsely Elevate Cortisol
- Glucocorticoids (stop day of testing)
- Spironolactone (stop day of testing)
- Estrogen-containing drugs (stop 4-6 weeks prior) 2, 1
Conditions Affecting CBG Levels
Low CBG (falsely low total cortisol):
- Cirrhosis
- Nephrotic syndrome
- Critical illness
High CBG (falsely high total cortisol):
- Estrogen therapy
- Pregnancy
Consider measuring CBG levels concurrently if these conditions are present to ensure accurate interpretation of total cortisol values 1, 2.
Rare Conditions
- CBG deficiency
- Glucocorticoid resistance
- Glucocorticoid hypersensitivity
These can severely affect test interpretation and require specialized evaluation 1.
Low-Dose (1 µg) Test
The low-dose test provides no better diagnostic accuracy than the 250 µg test for primary adrenal insufficiency 1. Reserve the 1 µg test only for situations when cosyntropin is in short supply 1. The low-dose test may have higher sensitivity for secondary adrenal insufficiency, but this is not the primary indication for ACTH testing 1.
Clinical Context
In acutely ill patients with severe symptoms or adrenal crisis (volume depletion, hypotension, hyponatremia, hyperkalemia, fever, abdominal pain, hyperpigmentation, or hypoglycemia in children), immediately administer IV hydrocortisone at stress doses without waiting for test results, as delayed treatment increases morbidity and mortality 1.