What is the best test for measuring testosterone levels?

Best Test for Measuring Testosterone Levels

The best test for measuring testosterone is early morning serum total testosterone, measured on two separate occasions, preferably using the same laboratory with consistent methods. 1

Primary Testing Approach

• Total testosterone should be measured in the early morning (before 11:00 AM) to account for the significant diurnal variation in testosterone production 1, 2
• Two separate measurements are required for diagnosis, as there can be significant intra-individual variability 1
• The threshold for low testosterone is defined as consistently <300 ng/dL on at least two measurements 1
• Testing should be performed using a reliable assay, preferably in a laboratory certified by an accuracy-based program such as the CDC's Hormone Standardization Program 3

When to Consider Additional Testing

• Free testosterone testing should be considered in specific situations:

  • When total testosterone levels are borderline (280-350 ng/dL) 4
  • In patients with obesity, as they often have low sex hormone-binding globulin (SHBG) levels 1
  • When there are suspected abnormalities in SHBG 1

• Free testosterone is best measured by equilibrium dialysis method, which is more accurate than calculated or analog methods 1, 3

  • Many local laboratories use analog methods which have limited reliability 1
  • Free testosterone index (total testosterone/SHBG ratio) can be used when equilibrium dialysis is not available 1

Follow-up Testing When Testosterone is Low

• Luteinizing hormone (LH) levels should be measured to determine if hypogonadism is primary (testicular) or secondary (pituitary/hypothalamic) 1
• Serum prolactin should be measured in patients with low testosterone and low/normal LH levels 1
• Estradiol should be measured in patients with breast symptoms or gynecomastia 1
• Men with total testosterone <150 ng/dL and low/normal LH should undergo pituitary MRI regardless of prolactin levels 1

Common Pitfalls and Caveats

• Laboratory reference ranges for testosterone vary widely between institutions, with low reference values ranging from 130 to 450 ng/dL (350% difference) 5
• Total testosterone alone has limited specificity for diagnosing hypogonadism except when levels are very low (<150 ng/dL) 4
• Total testosterone must exceed 350-400 ng/dL to reliably predict normal free testosterone 4
• Screening questionnaires should not be used as a substitute for laboratory testing due to variable sensitivity and specificity 1
• Testosterone levels should be measured in a fasting state for optimal accuracy 3

Special Populations

• In obese men, morning free testosterone by equilibrium dialysis is essential, as obesity can lower SHBG levels while free testosterone may remain normal 1
• Testosterone testing should be considered in patients with unexplained anemia, bone density loss, diabetes, exposure to chemotherapy, testicular radiation, HIV/AIDS, chronic narcotic use, male infertility, pituitary dysfunction, and chronic corticosteroid use 1
• For HIV-infected men with symptoms of fatigue, weight loss, loss of libido, erectile dysfunction, or depressive symptoms, morning serum total testosterone measurements should be obtained 1