Cortisol assessment, in saliva and urine

First line
Requires a medical indication

Test to investigate possible high cortisol with two late-night saliva samples and two 24-hour urine collections. It combines two screening tests for Cushing's syndrome: one assesses the night-time fall and the other reflects the free cortisol excreted during the day. It requires a medical indication and four correctly collected samples.

Requires a medical prescription

Results in 5-7 working days

Saliva and 24-hour urine sample. Includes: Salivary cortisol · 24-hour urine free cortisol. See what each one measures

This is the appropriate assessment when a doctor has already seen reasons to suspect cortisol excess. If that suspicion does not yet exist, the likelihood that a high result is a false positive is very high, and each one opens an expensive and distressing investigation.

What it is for

It is used to investigate possible cortisol excess from two different angles. Saliva looks for loss of the night-time low and urine assesses daily production. When the results agree and fit the clinical picture, the assessment becomes more consistent.

It does not locate the source of the problem or automatically confirm Cushing's syndrome. Choosing this profile over a single test depends on the clinical suspicion and on the conditions that can affect each method.

When it is recommended

Medical suspicion of Cushing's syndrome that justifies using two screening methods
Previous borderline or conflicting results that need to be repeated and compared
Suspected cyclical hypercortisolism, in which repetition may be particularly relevant
It is not recommended as a general assessment of obesity, tiredness or stress without specific clinical signs

Clinical comment

Each method needs repeating because cortisol varies from one day to the next and is released in pulses. That is why this protocol includes two nights of saliva and two full days of urine. A single sample of each type would leave both tests incomplete.

Agreement does not remove the need for interpretation. Alcohol, acute illness, certain medicines, some mood disorders and intense exercise can alter both methods. Kidney function and the quality of the collection particularly affect urine; sleep schedule and contamination from corticosteroid creams affect saliva.

If the results do not match, it does not automatically mean that one of them is wrong. They may need repeating at another time, the collection may need reviewing, or a cyclical pattern may need to be considered with the endocrinologist leading the assessment.

Dr. Antonio Mas

What is tested

Salivary cortisol (two nights): Measures the free fraction of cortisol at the end of the waking period over two nights.
24-hour urine free cortisol (two collections): Reflects the amount of free cortisol excreted over two complete days.

How to prepare

For saliva, collect a sample at bedtime on two nights. In the 15 minutes beforehand, do not eat, drink, smoke or brush your teeth.

For urine, do two complete 24-hour collections. Discard the first void, note the time and keep all the following ones up to and including the first one the next day at the same time. Store the containers according to the partner centre's instructions.

Saliva and urine do not have to be done on the same days. Avoid alcohol and intense exercise, and report any corticosteroid, including inhalers, creams and injections.

You can add

Without another needle. It is offered when you order, so you do not have to decide now.

General blood testA general blood test with full blood count, glucose, cholesterol and kidney and liver function: the basic parameters included in any general check-up.+€59.00
General + nutritional blood testEverything in the general blood test (anaemia, kidneys, liver, cholesterol and glucose) plus iron stores, vitamin D, vitamin B12 and folic acid. It is the broadest add-on: it covers the general and the nutritional in one go.+€129.00

Related blood tests

We have other profiles in Other tests with more or fewer tests:

The most comprehensive is not always the most useful. This is the one we recommend to start with; the others cover specific situations.

What it is based on

Pituitary Society · Lancet Diabetes Endocrinol 9(12):847-875
Consensus on diagnosis and management of Cushing's disease: a guideline update
Recommends starting with dexamethasone suppression, urine free cortisol and/or late-night salivary cortisol, with high quality and a strong recommendation, so combining two of them is an option the consensus expressly allows. It is also what supports the four samples: two to three urine collections, because variability between samples reaches 50%, and at least two nights of saliva.
Endocrine Society · J Clin Endocrinol Metab 93(5):1526-1540
The Diagnosis of Cushing's Syndrome: An Endocrine Society Clinical Practice Guideline
Names the three screening tests with the number of samples written into them: at least two in urine and two in saliva. That is why one urine collection plus one night of saliva are not two tests, but two tests done by halves.

Protocol designed by Dr. Antonio Mas, specialist in Endocrinology and Nutrition, Spanish medical licence no. 080846289, and reviewed in September 2026.

This blood test is a standardised protocol and does not replace a medical consultation. If you have symptoms, a known condition or a complex clinical situation, speak to your doctor.