Last Updated: August 23, 2026
Cortisol testing is an important part of endocrine laboratory medicine because a single cortisol concentration cannot always be interpreted as simply “normal,” “high,” or “low.” The result must be evaluated together with the time of specimen collection, clinical circumstances, medication exposure, ACTH concentration, assay methodology, and the laboratory-specific reference interval.
This laboratory guide explains the cortisol blood test, normal cortisol range, morning cortisol, high cortisol, low cortisol, Cushing syndrome, Addison disease, ACTH stimulation testing, dexamethasone suppression testing, pre-analytical variables, analytical interference, and laboratory interpretation.
- What Is Cortisol?
- Functions of Cortisol
- HPA Axis and Cortisol Regulation
- What Is a Cortisol Blood Test?
- Why Is Cortisol Testing Ordered?
- Specimen Requirements
- Morning Cortisol
- Cortisol Normal Range
- High Cortisol Levels
- Low Cortisol Levels
- Cushing Syndrome
- Addison Disease
- Cortisol and ACTH Interpretation
- ACTH Stimulation Test
- Dexamethasone Suppression Test
- Urine and Salivary Cortisol
- Pre-Analytical Factors
- Analytical Methods and Interferences
- Laboratory Interpretation Algorithm
- Critical Clinical Situations
- Cortisol Blood Test Interpretation Table
- Common Laboratory Mistakes in Cortisol Testing
- Cortisol and Other Laboratory Tests
- Frequently Asked Questions
- Authoritative References
- Medical Disclaimer
- Conclusion
1. What Is Cortisol?
Cortisol is the principal glucocorticoid hormone produced by the adrenal cortex. The adrenal glands are located above the kidneys and contain an outer cortex and inner medulla. Cortisol is primarily synthesized in the zona fasciculata of the adrenal cortex.
Cortisol secretion is controlled through the hypothalamic-pituitary-adrenal, or HPA, axis. Cortisol plays important roles in glucose metabolism, cardiovascular function, inflammation, immune regulation, protein metabolism, fat metabolism, and the physiologic response to stress.
Because cortisol affects glucose availability, cortisol abnormalities may influence glucose measurements and metabolic status. For complementary laboratory interpretation, see the Blood Glucose Test guide.
2. Functions of Cortisol
Cortisol has widespread effects throughout the body. Major physiologic functions include:
- Supporting the normal stress response.
- Maintaining vascular tone and blood pressure.
- Promoting hepatic glucose production during fasting and stress.
- Influencing protein and lipid metabolism.
- Modulating inflammation and immune activity.
- Supporting cardiovascular function.
- Influencing bone metabolism.
- Helping maintain adequate energy availability during illness or physiologic stress.
Chronic cortisol excess can contribute to hyperglycemia and altered glucose metabolism. When long-term glycemic status is clinically relevant, the HbA1c Blood Test guide provides additional laboratory context.
3. HPA Axis and Cortisol Regulation
The hypothalamus releases corticotropin-releasing hormone (CRH), which stimulates the anterior pituitary gland to produce adrenocorticotropic hormone (ACTH). ACTH then stimulates the adrenal cortex to produce cortisol.
The simplified pathway is:
Cortisol then provides negative feedback to both the hypothalamus and pituitary. When circulating cortisol becomes high, CRH and ACTH secretion are normally reduced. When cortisol is low, ACTH may increase if pituitary function is intact.
Because pituitary and thyroid abnormalities may coexist or produce overlapping nonspecific symptoms such as fatigue and weakness, interpretation sometimes includes additional endocrine testing. See the Thyroid Function Tests laboratory guide.
4. What Is a Cortisol Blood Test?
A cortisol blood test measures cortisol concentration in serum or plasma. Depending on the clinical question, a clinician may order:
- 8 AM or early-morning serum cortisol.
- Afternoon cortisol.
- Cortisol together with plasma ACTH.
- Cortisol before and after cosyntropin stimulation.
- Cortisol after dexamethasone administration.
- Serial cortisol measurements.
Other specimens, especially 24-hour urinary free cortisol and late-night salivary cortisol, may be more appropriate for specific investigations such as suspected hypercortisolism.
5. Why Is Cortisol Testing Ordered?
Cortisol testing may be requested when there is clinical concern for:
- Adrenal insufficiency.
- Addison disease.
- Cushing syndrome.
- Pituitary dysfunction.
- Adrenal tumors.
- ACTH-producing tumors.
- Suppression of the HPA axis after glucocorticoid exposure.
- Unexplained hypotension or electrolyte abnormalities.
- Evaluation of adrenal reserve.
Because many cortisol-related symptoms are nonspecific, clinicians frequently interpret cortisol alongside a broader laboratory profile such as electrolytes, renal markers, glucose, CBC, and inflammatory markers. For blood-cell evaluation, review the Complete Blood Count (CBC) guide.
6. Specimen Requirements for Cortisol Blood Testing
| Parameter | Laboratory Consideration |
|---|---|
| Specimen | Serum or plasma depending on analyzer and laboratory procedure. |
| Collection time | Must be documented because cortisol demonstrates marked diurnal variation. |
| Morning sample | Often collected around 8 AM when physiologic cortisol is expected to be relatively high. |
| Patient condition | Stress, pain, exercise, acute illness and sleep disruption can influence cortisol. |
| Medication history | Glucocorticoids, estrogen-containing medication and several other drugs can affect interpretation. |
| Dynamic testing | Exact collection times must follow the stimulation or suppression protocol. |
7. Morning Cortisol: Why Timing Matters
Cortisol follows a circadian rhythm. Concentrations are generally highest in the early morning and decline throughout the day, reaching relatively low levels around late evening or nighttime.
For this reason, an 8 AM cortisol cannot be interpreted using an evening reference interval, and an evening cortisol should not be compared with an early-morning range.
Morning cortisol is particularly useful during the initial evaluation of suspected adrenal insufficiency. Extremely low values may increase suspicion for adrenal failure, whereas clearly adequate values may make significant adrenal insufficiency less likely. Intermediate values frequently require dynamic testing.
8. Cortisol Normal Range
A commonly cited example for blood collected at approximately 8 AM is:
| Sample | Example Reference Range | SI Units |
|---|---|---|
| Morning blood cortisol, approximately 8 AM | 5–25 µg/dL | 138–690 nmol/L |
This range is an educational example and must not replace the reference interval printed on the patient's laboratory report. Cortisol assays are not completely interchangeable, and intervals vary according to analytical method, laboratory population, collection time, specimen type, and clinical context.
How to Convert Cortisol Units
Cortisol results are commonly reported as µg/dL or nmol/L.
1 µg/dL cortisol ≈ 27.6 nmol/L
Therefore, a cortisol result should never be labeled abnormal based only on a value found on the internet. The correct comparator is the assay-specific laboratory reference interval.
9. High Cortisol Levels
An elevated cortisol concentration may result from true endogenous cortisol excess or from physiologic, pharmacologic, and analytical factors.
Possible Causes of High Cortisol
- Cushing syndrome.
- ACTH-producing pituitary disease.
- Ectopic ACTH production.
- Cortisol-producing adrenal tumor.
- Severe physical or psychological stress.
- Acute illness.
- Major surgery or trauma.
- Estrogen-associated increases in cortisol-binding globulin.
- Pregnancy-related physiologic changes.
- Certain medications.
- Assay interference.
Inflammation and acute illness may substantially change endocrine physiology. When inflammatory markers are being assessed at the same time, the High CRP Levels laboratory guide can help provide broader laboratory context.
Symptoms Associated With Chronic Cortisol Excess
Depending on the cause and severity, chronic cortisol excess may be associated with:
- Central weight gain.
- Rounded facial appearance.
- Proximal muscle weakness.
- Easy bruising.
- Wide purple striae.
- Hypertension.
- Hyperglycemia.
- Reduced bone density.
- Menstrual abnormalities.
- Mood changes.
These findings are not specific enough to diagnose Cushing syndrome without appropriate biochemical testing.
10. Low Cortisol Levels
Low cortisol may occur because the adrenal glands cannot produce adequate cortisol or because ACTH stimulation from the pituitary is insufficient.
Possible Causes
- Primary adrenal insufficiency.
- Addison disease.
- Secondary adrenal insufficiency due to pituitary disease.
- Tertiary adrenal suppression related to hypothalamic dysfunction.
- Suppression after prolonged glucocorticoid exposure.
- Sudden withdrawal of chronic glucocorticoid therapy.
- Adrenal destruction, hemorrhage, infection or infiltrative disease.
- Selected congenital adrenal disorders.
Possible Symptoms
- Persistent fatigue.
- Weakness.
- Loss of appetite.
- Weight loss.
- Abdominal symptoms.
- Low blood pressure.
- Dizziness.
- Electrolyte abnormalities.
Low cortisol should always be evaluated in clinical context. Kidney function and electrolyte abnormalities may also be relevant in acutely ill patients. For additional laboratory interpretation, see the Creatinine and eGFR laboratory guide.
11. Cortisol Testing and Cushing Syndrome
Cushing syndrome describes the clinical consequences of prolonged exposure to excessive glucocorticoid activity. The most common overall cause is exposure to exogenous glucocorticoid medication.
Endogenous Cushing syndrome may arise from:
- Pituitary ACTH excess.
- Ectopic ACTH production.
- ACTH-independent adrenal cortisol production.
Recommended Initial Biochemical Approaches
Commonly recommended tests include:
- 24-hour urinary free cortisol (UFC), usually with repeat collection.
- Late-night salivary cortisol, generally with more than one measurement.
- 1-mg overnight dexamethasone suppression test.
- Longer low-dose dexamethasone suppression testing in selected circumstances.
Cushing Syndrome vs Cushing Disease
The terms should not be used interchangeably. Cushing syndrome describes cortisol excess regardless of cause. Cushing disease specifically refers to pituitary ACTH-dependent Cushing syndrome.
12. Cortisol Testing and Addison Disease
Addison disease is primary adrenal insufficiency, meaning that the adrenal cortex cannot produce adequate glucocorticoid hormones and, frequently, mineralocorticoids.
Laboratory findings may include:
- Low cortisol.
- Elevated ACTH in primary adrenal insufficiency.
- Hyponatremia.
- Hyperkalemia in mineralocorticoid deficiency.
- Hypoglycemia in some patients.
- Elevated renin and reduced aldosterone in appropriate cases.
Because nutritional deficiency may also produce weakness and fatigue, additional tests may sometimes be performed depending on the clinical presentation. See the Vitamin B12 Blood Test guide and Vitamin D Blood Test guide.
13. Cortisol and ACTH: Laboratory Interpretation
Paired cortisol and ACTH measurements may help identify the level of dysfunction within the HPA axis.
| Cortisol | ACTH | Possible Interpretation |
|---|---|---|
| Low | High | Pattern may support primary adrenal insufficiency. |
| Low | Low or inappropriately normal | May suggest secondary or tertiary adrenal insufficiency. |
| High | High or inappropriately normal | May occur with ACTH-dependent hypercortisolism. |
| High | Suppressed | May suggest ACTH-independent adrenal cortisol production. |
These patterns are simplified teaching models and do not replace formal endocrine investigation.
ACTH Specimen Handling
ACTH is more pre-analytically demanding than serum cortisol. Collection tubes, temperature requirements, processing time, centrifugation, and storage must follow the laboratory's validated protocol and analyzer manufacturer's requirements.
14. ACTH Stimulation Test
The ACTH stimulation test, commonly performed using cosyntropin, evaluates adrenal cortisol response to ACTH stimulation.
A typical protocol includes:
- Collection of a baseline cortisol sample.
- Administration of synthetic ACTH according to the approved protocol.
- Collection of cortisol at specified post-stimulation times, commonly 30 and/or 60 minutes depending on protocol.
- Comparison with an assay-specific stimulated cortisol cutoff.
An Endocrine Society guideline historically described peak cortisol below approximately 18 µg/dL (500 nmol/L) after stimulation as suggestive of adrenal insufficiency, while explicitly recognizing that the threshold is assay dependent. Modern assays may require different cutoffs.
15. Dexamethasone Suppression Test
The dexamethasone suppression test evaluates whether administered dexamethasone appropriately suppresses endogenous ACTH and cortisol production.
1-mg Overnight Dexamethasone Suppression Test
In a commonly used protocol, dexamethasone is administered late in the evening, followed by serum cortisol measurement the following morning.
Failure of cortisol to suppress appropriately may raise suspicion for autonomous or pathologic cortisol production, although false-positive results can occur.
Factors Affecting Dexamethasone Testing
- Medication interactions that alter dexamethasone metabolism.
- Non-adherence or incorrect dosing time.
- Estrogen-associated changes in cortisol-binding globulin.
- Acute illness.
- Alcohol-related physiologic effects.
- Severe stress.
- Assay-specific factors.
16. Serum vs Urinary vs Salivary Cortisol
| Test | Main Laboratory Role | Important Limitation |
|---|---|---|
| Serum cortisol | Morning assessment, dynamic stimulation or suppression testing. | Influenced by circadian rhythm and cortisol-binding proteins. |
| 24-hour urinary free cortisol | Assessment of integrated free cortisol excretion; useful in suspected hypercortisolism. | Requires accurate 24-hour urine collection. |
| Late-night salivary cortisol | Assesses loss of the normal late-night cortisol nadir. | Collection timing, contamination and sleep pattern matter. |
When evaluating a 24-hour urine specimen, completeness of collection is essential. For a broader overview of laboratory urine analysis and specimen considerations, see the Urinalysis Complete Guide.
17. Pre-Analytical Factors Affecting Cortisol
Pre-analytical variables are particularly important because cortisol responds rapidly to physiologic conditions.
Collection Time
The collection time should be recorded accurately. A cortisol result without a reliable collection time loses considerable interpretive value.
Stress
Venipuncture anxiety, pain, hospitalization, acute illness, surgery and trauma may increase cortisol.
Exercise
Vigorous physical activity can alter cortisol. Patient preparation should follow the ordering clinician's and laboratory's instructions.
Sleep Pattern and Shift Work
Normal circadian assumptions may not apply perfectly to people with abnormal sleep schedules or night-shift work.
Medications
Medication history is essential. Glucocorticoid exposure may come from oral tablets, injections, inhalers, topical formulations, nasal preparations and other routes.
Patients should not stop prescribed steroids independently before cortisol testing. Abrupt discontinuation may be dangerous.
Estrogen and Cortisol-Binding Globulin
Most circulating cortisol is protein bound. Increased cortisol-binding globulin can raise measured total serum cortisol even when biologically active free cortisol is not increased to the same extent.
18. Analytical Methods and Laboratory Interferences
Cortisol is commonly measured using automated immunoassays, while liquid chromatography-tandem mass spectrometry (LC-MS/MS) may be used in specialized settings.
Immunoassays
Advantages include:
- High throughput.
- Automation.
- Rapid turnaround.
- Wide availability in routine clinical laboratories.
Potential limitations include:
- Cross-reactivity with structurally related steroids.
- Drug or metabolite interference.
- Method-to-method bias.
- Assay-specific calibration differences.
LC-MS/MS
LC-MS/MS can offer greater analytical specificity and the ability to distinguish cortisol from related steroid compounds, although availability, workflow complexity and cost may limit routine use.
Hemolysis, Icterus and Lipemia
The effect of hemolysis, icterus or lipemia depends on the analytical platform. Laboratories should use manufacturer interference studies and their own validated procedures. For a detailed discussion of specimen interference, see Hemolysis in Clinical Chemistry.
Quality Control
Cortisol patient results should only be reported when analytical performance is acceptable according to the laboratory quality management system. Internal QC should be reviewed for shifts, trends, random errors and systematic errors.
For a detailed overview of Levey-Jennings charts and Westgard rules, see the Internal Quality Control (IQC) guide.
19. Step-by-Step Laboratory Interpretation of Cortisol
A systematic laboratory approach can reduce misinterpretation.
Step 1: Confirm the Clinical Question
Determine whether the test is investigating:
- Possible adrenal insufficiency.
- Possible cortisol excess.
- A dynamic stimulation test.
- A suppression test.
- Monitoring of a known endocrine disorder.
Step 2: Verify Specimen Collection Time
Morning and evening results are not interchangeable.
Step 3: Check the Reference Interval
Use the laboratory's own method- and time-specific interval.
Step 4: Review Medications
Ask about systemic, inhaled, topical and injected glucocorticoids as well as medications affecting steroid metabolism or binding proteins.
Step 5: Review Patient Condition
Consider acute illness, hospitalization, surgery, psychological stress, sleep disruption, exercise, pregnancy and estrogen exposure.
Step 6: Review Related Laboratory Results
Potentially useful associated measurements include ACTH, sodium, potassium, glucose, creatinine, CBC, thyroid tests and liver tests.
When hepatic abnormalities are present, the High Liver Enzymes ALT & AST guide provides additional interpretation of common liver markers.
Step 7: Determine Whether Dynamic Testing Is Needed
An isolated borderline cortisol should not be overinterpreted. Depending on the clinical question, ACTH stimulation, dexamethasone suppression, urinary free cortisol or late-night salivary cortisol may provide more diagnostically useful information.
20. Critical Clinical Situations
Adrenal Crisis
Acute adrenal crisis is a potentially life-threatening condition. Possible findings include severe weakness, hypotension, vomiting, abdominal symptoms, altered mental status, electrolyte abnormalities and hypoglycemia.
Acute Illness and Cortisol
Severe illness activates the HPA axis and may substantially alter cortisol secretion, binding proteins and metabolism. Consequently, interpretation in critical illness can be more complex than outpatient interpretation.
21. Cortisol Blood Test Interpretation Table
| Finding | Possible Explanation | Common Next Consideration |
|---|---|---|
| Low morning cortisol | Adrenal insufficiency, HPA suppression or pituitary disease | ACTH measurement and/or stimulation testing depending on context |
| High morning cortisol | Stress, acute illness, increased binding protein or cortisol excess | Clinical assessment; dedicated Cushing testing if appropriate |
| High cortisol + high ACTH | Possible ACTH-dependent cortisol excess | Endocrine evaluation after biochemical confirmation |
| High cortisol + suppressed ACTH | Possible ACTH-independent cortisol production | Further endocrine investigation |
| Low cortisol + high ACTH | Possible primary adrenal insufficiency | Confirmatory adrenal evaluation |
| Low cortisol + low/normal ACTH | Possible central adrenal insufficiency | Pituitary/HPA axis evaluation |
22. Common Laboratory Mistakes in Cortisol Testing
- Interpreting a random cortisol using a morning reference range.
- Ignoring specimen collection time.
- Using one serum cortisol result to diagnose Cushing syndrome.
- Ignoring current or recent glucocorticoid exposure.
- Applying an old ACTH stimulation cutoff to a newer assay without validation.
- Ignoring cortisol-binding protein effects.
- Failing to follow precise dynamic-test collection times.
- Interpreting abnormal results without clinical context.
- Reporting patient results when QC performance is unacceptable.
23. Cortisol and Other Laboratory Tests
Cortisol abnormalities rarely exist in a laboratory vacuum. Depending on the clinical presentation, additional testing may include:
- ACTH.
- Sodium and potassium.
- Glucose.
- HbA1c.
- Renal function.
- Liver function.
- Thyroid function.
- CBC.
- Renin and aldosterone.
Interpretation of cortisol should therefore focus on the complete biochemical and clinical pattern rather than a single numeric result.
24. Frequently Asked Questions
What is a normal cortisol level in the morning?
One commonly cited reference example for an approximately 8 AM serum cortisol is 5–25 µg/dL (138–690 nmol/L). However, the reference interval varies by laboratory and analytical method, so the range printed on the laboratory report should be used.
What time should a cortisol blood test be done?
Morning cortisol is commonly collected around 8 AM because cortisol is normally relatively high early in the day. Other collection times may be specifically requested depending on the diagnostic protocol.
What does high cortisol mean?
High cortisol may occur with Cushing syndrome, acute illness, physiologic stress, some medications, altered cortisol-binding proteins or other conditions. A single high result does not establish Cushing syndrome.
What does low cortisol mean?
Low cortisol may occur with primary adrenal insufficiency, pituitary or hypothalamic disease, or suppression of the HPA axis after glucocorticoid exposure. The result must be interpreted according to collection time and clinical context.
Can stress increase cortisol?
Yes. Physical and psychological stress can activate the HPA axis and increase cortisol secretion. Acute illness, surgery, trauma and strenuous exercise can also alter cortisol concentrations.
Can a cortisol blood test diagnose Cushing syndrome?
A random serum cortisol concentration alone should not be used to diagnose Cushing syndrome. Appropriate screening tests include late-night salivary cortisol, urinary free cortisol and dexamethasone suppression testing depending on the patient.
Can one low cortisol result diagnose Addison disease?
Usually not by itself. Morning cortisol may help identify patients who require further investigation, while ACTH measurement and ACTH stimulation testing are commonly used during evaluation.
What is the difference between Addison disease and adrenal insufficiency?
Addison disease refers specifically to primary adrenal insufficiency caused by failure of the adrenal cortex. Adrenal insufficiency can also be secondary or tertiary when the problem originates in the pituitary or hypothalamic-HPA axis.
Can steroid medications affect cortisol test results?
Yes. Glucocorticoids can suppress the HPA axis, and some synthetic steroids or metabolites may also interfere with certain laboratory methods. Patients should tell their clinician and laboratory about all steroid medications and should not stop prescribed therapy without medical guidance.
Is cortisol higher in the morning or at night?
In people with a conventional sleep-wake cycle, cortisol is normally higher in the morning and falls during the day, with a relatively low concentration late at night.
25. Authoritative Medical References
- MedlinePlus — Cortisol Test.
- Endocrine Society — Diagnosis of Cushing’s Syndrome Clinical Practice Guideline.
- Endocrine Society — Primary Adrenal Insufficiency Clinical Practice Guideline.
- Endocrine Society — Hormone Replacement in Hypopituitarism Guideline (Central Adrenal Insufficiency).
- Endocrine Society — Cushing’s Syndrome and Cushing Disease.
- NIDDK — Cushing’s Syndrome.
- NIDDK — Adrenal Insufficiency & Addison’s Disease.
Reference ranges and diagnostic thresholds can change according to assay methodology. Laboratories should follow their validated method, manufacturer instructions, current professional guidance and local clinical protocols.
26. Medical Disclaimer
Conclusion
The cortisol blood test is a valuable laboratory tool for evaluating adrenal and pituitary function, but interpretation requires much more than comparing a number with a generic reference range. Cortisol follows a strong circadian rhythm, responds to physiologic stress, is affected by medications and binding proteins, and varies between analytical methods.
A morning cortisol can provide useful initial information when adrenal insufficiency is suspected, but indeterminate results often require ACTH measurement or dynamic stimulation testing. Conversely, random serum cortisol is not an appropriate stand-alone screening test for Cushing syndrome; urinary free cortisol, late-night salivary cortisol and dexamethasone suppression testing are more appropriate in selected patients.
For laboratory professionals, accurate collection timing, correct specimen handling, appropriate quality control, awareness of assay interference and use of method-specific reference intervals are essential for reliable cortisol reporting and clinical interpretation.