Hormones are not steady numbers. Several of them are released in a daily rhythm or in pulses, so the value in the tube depends on the hour it was collected. A cortisol of 6 mcg/dL means something different at 8 am and at 10 pm. A testosterone drawn after lunch can read a third lower than the same man's morning value. Sampling time is not an administrative detail; it is a condition of interpreting the result.

Cortisol through the day

Cortisol has the strongest daily rhythm of the common endocrine tests. It rises before waking, peaks in the first hour after waking, then falls to its lowest point around midnight. The Endocrine Society guideline on primary adrenal insufficiency uses a morning cortisol below 5 mcg/dL, which is 140 nmol/L, alongside ACTH as a preliminary sign of adrenal insufficiency. The guideline on Cushing syndrome uses the opposite end of the rhythm and asks for two late-night salivary cortisol measurements, because losing the night-time trough is an early feature of cortisol excess.

Testosterone and the morning rule

Testosterone falls through the day and drops after eating. The 2018 Endocrine Society guideline states that clinicians should measure total testosterone concentrations on two separate mornings when the patient is fasting, using an assay standardised against a programme such as the CDC Hormone Standardization Program. A single afternoon testosterone is the most common reason men are told they have low testosterone when they do not, and the most common reason a genuine deficiency is dismissed.

Cycle day and pituitary hormones

In women who still menstruate, oestradiol, progesterone, FSH and LH move through predictable phases, so a value cannot be read without knowing the cycle day it was drawn on. Progesterone is measured in the luteal phase when the question is ovulation. Prolactin is best drawn three to four hours after waking, according to MedlinePlus, and rises with sleep, stress and breast stimulation, so a mildly high prolactin from a difficult venepuncture is usually repeated before anyone orders imaging.

Hormone	When to draw	Why
Cortisol (deficiency question)	Early morning	Peak value; below 5 mcg/dL is suggestive
Cortisol (excess question)	Late night, salivary, two samples	Loss of the night-time trough
Total testosterone	Two separate fasting mornings	2018 Endocrine Society guideline
Prolactin	Three to four hours after waking	Falls through the day, rises with stress
Oestradiol, progesterone, FSH	By cycle day for the question asked	Values shift across the cycle

When you book hormone testing, ask what time the sample should be taken and, for cycle hormones, which day. Write the collection time on your own copy of the result. If a hormone value is going to change a prescription, it is worth taking twice under the same conditions before anyone acts on it.

TSH also has a diurnal pattern, with a nocturnal peak and lower values in the late morning and afternoon. The variation is usually modest relative to treatment decisions, but it is enough to explain why a patient monitored at 8 am one year and 3 pm the next appears to have shifted. Keeping the collection time consistent, and keeping levothyroxine dosing consistent relative to the draw, removes that noise.

Pulsatile secretion is a separate issue from diurnal rhythm. LH is secreted in pulses roughly every 60 to 90 minutes, so single values are interpreted as a pattern with FSH and oestradiol rather than alone. Growth hormone is pulsatile enough that random measurement is uninterpretable, which is why IGF-1 and dynamic testing are used instead.

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Cortisol through the day

Testosterone and the morning rule

Cycle day and pituitary hormones

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