Test Catalog

Test ID: CORTO    
Cortisol, Free and Total, Serum

Useful For Suggests clinical disorders or settings where the test may be helpful

Assessment of cortisol status in cases where there is known or a suspected abnormality in cortisol-binding proteins or albumin


Assessment of adrenal function in the critically ill or stressed patient, thus preventing unnecessary use of glucocorticoid therapy


Second-order testing when cortisol measurement by immunoassay (eg, CORT / Cortisol, Serum) gives results that are not consistent with clinical symptoms, or if patients are known to, or suspected of, taking exogenous synthetic steroids


An adjunct in the differential diagnosis of primary and secondary adrenal insufficiency


An adjunct in the differential diagnosis of Cushing syndrome

Clinical Information Discusses physiology, pathophysiology, and general clinical aspects, as they relate to a laboratory test

Cortisol, the main glucocorticoid (representing 75%-95% of the plasma corticoids), plays a critical role in glucose metabolism and in the body's response to stress. Both hypercortisolism (Cushing disease) and hypocortisolism (Addison disease) can cause disease. Cortisol is also used to treat skin disease, allergic disorders, respiratory system disease, inflammatory disorders, and nephrotic syndrome.


Cortisol levels are regulated by adrenocorticotropic hormone (ACTH), which is synthesized by the pituitary in response to corticotropin-releasing hormone (CRH). CRH is released in a cyclic fashion by the hypothalamus, resulting in diurnal peaks (6-8 a.m.) and nadirs (11 p.m.) in plasma ACTH and cortisol levels.


The majority of cortisol circulates bound to corticosteroid-binding globulin (CBG) and albumin. Normally, less than 5% of circulating cortisol is free (unbound). Only free cortisol can access the enzyme transporters in liver, kidney, and other tissues that mediate metabolic and excretory clearance.


Historically, measurements of free cortisol have been achieved from indirect means using a ratio known as the free cortisol index. This measurement takes into account the amount of total cortisol and CBG to give a percentage and, ultimately, absolute value of free cortisol. These methods do not take into account the possibility variations in albumin levels. These calculations also rely on CBG, which can be lowered in critically ill patients despite normal adrenal function. Equilibrium dialysis best serves to separate free from bound cortisol without disrupting the bound fraction.


Pathological hypercortisolism due to endogenous or exogenous glucocorticoids is termed Cushing syndrome. Signs and symptoms of pathological hypercortisolism may include central obesity, hypertension, hyperglycemia, hirsutism, muscle weakness, and osteoporosis. However, these symptoms and signs are not specific for pathological hypercortisolism. The majority of individuals with some or all of the symptoms and signs will not suffer from Cushing syndrome.


When Cushing syndrome is present, the most common cause is iatrogenic, due to repeated or prolonged administration of, mostly, synthetic corticosteroids. Spontaneous Cushing syndrome is less common and results from either primary adrenal disease (adenoma, carcinoma, or nodular hyperplasia) or an excess of ACTH (from a pituitary tumor or an ectopic source). ACTH-dependent Cushing syndrome due to a pituitary corticotroph adenoma is the most frequently diagnosed subtype, most commonly seen in women in the third through fifth decades of life. The onset is insidious and usually occurs 2 to 5 years before a clinical diagnosis is made.


Hypocortisolism most commonly presents with nonspecific lassitude, weakness, hypotension, and weight loss. Depending on the cause, hyperpigmentation may be present. More advanced cases and patients submitted to physical stress (ie, infection, spontaneous or surgical trauma) also may present with abdominal pain, hyponatremia, hyperkalemia, hypoglycemia, and in extreme cases, cardiovascular shock and renal failure.


The more common causes of hypocortisolism are:

Primary adrenal insufficiency:

-Addison disease

-Congenital adrenal hyperplasia, defects in enzymes involved in cortisol synthesis


Secondary adrenal insufficiency:

-Prior, prolonged corticosteroid therapy

-Pituitary insufficiency

-Hypothalamic insufficiency


See Steroid Pathways in Special Instructions.

Reference Values Describes reference intervals and additional information for interpretation of test results. May include intervals based on age and sex when appropriate. Intervals are Mayo-derived, unless otherwise designated. If an interpretive report is provided, the reference value field will state this.


6-10:30 a.m. Collection: 0.121-1.065 mcg/dL



a.pm: 5-25 mcg/dL

p.m.: 2-14 mcg/dL

Interpretation Provides information to assist in interpretation of the test results

Cortisol is converted to cortisone in human kidneys and cortisone is less active toward the mineralcorticoid receptor. The conversion of cortisol to cortisone in the kidney is mediated by 11B-hydroxysteroid dehydrogenase isoform-2. Also, cortisol renal clearance will be reduced when there is a deficiency in the cytochrome P450 3A5 (CYP3A5) enzyme as well as a deficiency in P-glycoprotein.


Cortisol-binding globulin (CBG) has a low capacity and high affinity for cortisol, whereas albumin has a high capacity and low affinity for binding cortisol. Variations in CBG and serum albumin due to renal or liver disease may have a major impact on free cortisol.


Based on the study by Bancos(1), normal ranges of free cortisol found in patients without adrenal insufficiency were:

-Free cortisol at baseline: median 0.400 mcg/dL (interquartile range: IQR 2.5-97.5% - 0.110-1.425 mcg/dL)

-Free cortisol at 30 minutes: median 1.355 mcg/dL (IQR 2.5-97.5% - 0.885-2.440 mcg/dL)

-Free cortisol at 60 minutes: median 1.720 mcg/dL (IQR 2.5-97.5% - 1.230-2.930 mcg/dL)


Based on the study by Bancos,(1) the following cutoffs were calculated for exclusion of adrenal insufficiency:

-Free cortisol at baseline*: greater than 0.271 mcg/dL (>271 ng/dL, area under the curve: AUC 0.81)

-Free cortisol at 30 minutes: greater than 0.873 mcg/dL (>873 ng/dL, AUC 0.99)

-Free cortisol at 60 minutes: greater than 1.190 mcg/dL (>1,190 ng/dL, AUC 0.99)

(*please note that baseline free cortisol should not be used to exclude adrenal insufficiency given low performance)


The use of free cortisol in the management of glucocorticoid levels in the stressed patient due to major surgery or trauma requires further studies to establish clinical dosing levels and efficacy.


Cortisol pediatric reference ranges are generally the same as adults as confirmed by peer-reviewed literature.(2)


In primary adrenal insufficiency, adrenocorticotropic hormone (ACTH) levels are increased and cortisol levels are decreased; in secondary adrenal insufficiency both ACTH and cortisol levels are decreased.


When symptoms of glucocorticoid deficiency are present and the 8 a.m. plasma cortisol value is less than 10 mcg/dL (or the 24-hour urinary free cortisol value is <50 mcg/24 hours), further studies are needed to establish the diagnosis. The 3 most frequently used tests are the ACTH (cosyntropin) stimulation test, the metyrapone test, and insulin-induced hypoglycemia test. First, the basal plasma ACTH concentration should be measured and the short cosyntropin stimulation test performed.


Symptoms or signs of Cushing syndrome in a patient with low serum and urine cortisol levels suggest possible exogenous synthetic steroid effects.

Cautions Discusses conditions that may cause diagnostic confusion, including improper specimen collection and handling, inappropriate test selection, and interfering substances

Cortisol levels may be increased in pregnancy and with exogenous estrogens. Use of the antineoplastic drug Mitotane also increases cortisol-binding globulin and total cortisol.


When cortisol assays are used for serial monitoring, the same methodology should be used throughout.


There is little, if any, value in an isolated p.m. serum cortisol measurement.


Acute stress (including hospitalization and surgery), alcoholism, depression, and many drugs (ie, exogenous cortisones, anticonvulsants) can obliterate normal diurnal variation, affect response to suppression/stimulation tests, and cause elevated baseline levels.

Clinical Reference Recommendations for in-depth reading of a clinical nature

1. Bancos I, Erickson D, Bryant S, et al: Performance of free versus total cortisol following cosyntropin stimulation testing in an outpatient setting. Endocr Pract 2015 Dec;21(12):1353-1363 doi: 10.4158/EP15820

2. Petersen KE: ACTH in normal children and children with pituitary and adrenal diseases. I. Measurement in plasma by radioimmunoassay-basal values. Acta Paediatr Scand 1981;70:341-345

3. Hamrahian AH, Oseni TS, Arafah BM: Measurements of serum free cortisol in critically ill patients. N Engl J Med 2004;350;16:1629-1638

4. Ho JT, Al-Musalhi H, Chapman MJ, et al: Septic shock and sepsis: a comparison of total and free plasma cortisol levels. J Clin Endocrinol Metab 2006;91:105-114

5. le Roux CW, Chapman GA, Kong WM, et al: Free cortisol index is better than serum total cortisol in determining hypothalamic-pituitary-adrenal status in patients undergoing surgery. J Clin Endocrinol Metab 2003;88:2045-2048

6. Huang W, Kalhorn TF, Baillie M, et al: Determination of free and total cortisol in plasma and urine by liquid chromatography-tandem mass spectrometry. Ther Drug Monit 2007;29(2):215-224

Special Instructions Library of PDFs including pertinent information and forms related to the test