Optimal Timing for Testosterone Testing
Draw testosterone levels between 8 AM and 10 AM for all men, regardless of age, to ensure accurate diagnosis of hypogonadism. 1, 2
Evidence-Based Timing Recommendations
Standard Morning Collection Window
The American Urological Association and Endocrine Society mandate that testosterone be measured between 8 AM and 10 AM on at least two separate mornings to account for diurnal variation and confirm persistently low levels. 1, 2, 3
Morning testosterone levels are 20-25% higher at 8 AM compared to 4 PM in men aged 30-40 years, with this difference declining to approximately 10% by age 70. 4
Two separate morning measurements are required because single measurements are insufficient due to significant intra-individual variability and assay differences. 1, 5
The Age-Related Controversy
While some older studies suggested that men over 45 years could be tested later in the day because diurnal variation diminishes with age 6, 7, 8, current guidelines from the American Urological Association and Endocrine Society do not make age-based exceptions and recommend 8-10 AM testing for all adult men. 1, 2
Even in men aged 66-80 years, 4 of 9 men with low afternoon testosterone had normal levels when tested before noon, demonstrating that clinically significant diurnal variation persists into older age. 4
The risk of false-positive hypogonadism diagnoses when testing outside morning hours justifies the universal 8-10 AM recommendation, as 17 men with at least one low afternoon measurement had normal testosterone at all three morning visits. 4
Additional Testing Considerations
Fasting Status
- Fasting is not required for testosterone measurement according to the American Urological Association, though some experts recommend it for consistency. 2, 5
Free Testosterone Timing
Free testosterone and bioavailable testosterone show greater diurnal variation than total testosterone in men under 75 years, making early morning collection (8-10 AM) even more critical when measuring these parameters. 6
Free testosterone should be measured by equilibrium dialysis or calculated using validated formulas when total testosterone is borderline (280-400 ng/dL) or when SHBG abnormalities are suspected. 2, 3, 5
Laboratory Consistency
Use the same laboratory and same methodology for serial measurements to minimize assay-related variability. 2
Ideally, testosterone should be measured using liquid chromatography tandem mass spectrometry (LC-MS/MS) in a laboratory certified by an accuracy-based benchmark. 5
Common Pitfalls to Avoid
Never diagnose hypogonadism based on a single testosterone measurement—two separate morning values below 300 ng/dL are required. 1, 2, 3
Do not test testosterone outside the 8-10 AM window, as this increases the risk of false-positive diagnoses, particularly in younger men. 1, 4
Avoid testing men who are acutely ill or under significant stress, as these conditions can transiently suppress testosterone levels. 1
Do not rely on afternoon or late-morning draws even in older men, as current guidelines do not support age-based exceptions to the morning collection requirement. 1, 2