What blood tests are recommended for the workup of low testosterone (hypogonadism) in men?

Blood Tests for Low Testosterone Workup in Men

Confirm the diagnosis with two separate early morning total testosterone measurements below 300 ng/dL using the same laboratory and methodology, then measure serum luteinizing hormone (LH) in all patients with confirmed low testosterone. 1

Initial Diagnostic Testing

Total Testosterone Measurement

• Obtain two separate fasting morning total testosterone levels (ideally between 7-11 AM) to confirm the diagnosis 1, 2
• Use a reliable assay, preferably liquid chromatography tandem mass spectrometry (LC-MS/MS) in a laboratory certified by an accuracy-based benchmark 3
• Both measurements should be below 300 ng/dL before proceeding with further workup 1, 2
• Use the same laboratory with identical methodology for both measurements to minimize variability 1

Free or Bioavailable Testosterone

• Measure free testosterone (by equilibrium dialysis) or calculate it using total testosterone, sex hormone-binding globulin (SHBG), and albumin when: 1
  • Total testosterone is near the lower limit of normal (close to 300 ng/dL)
  • SHBG abnormalities are suspected (obesity, aging, diabetes, thyroid disorders)
  • There is discordance between symptoms and total testosterone levels

Mandatory Secondary Testing

Luteinizing Hormone (LH)

• Measure serum LH in all patients with confirmed low testosterone to distinguish primary from secondary hypogonadism 1, 2
• Low or low-normal LH with low testosterone indicates secondary (central) hypogonadism and triggers additional testing 1, 4
• Elevated LH with low testosterone indicates primary (testicular) hypogonadism 1

Prolactin

• Measure serum prolactin immediately in all patients with low testosterone combined with low or low-normal LH levels 1, 4, 2
• If prolactin is elevated, repeat the measurement to exclude spurious elevation 1
• Persistently elevated prolactin requires referral to endocrinology and evaluation for prolactinoma 1, 4

Conditional Advanced Testing

Pituitary MRI

• Order pituitary MRI for men with: 1, 4
  • Total testosterone <150 ng/dL combined with low or low-normal LH, regardless of prolactin level
  • Persistently elevated prolactin levels
  • This identifies non-secreting pituitary adenomas that may not elevate prolactin

Additional Pituitary Hormones

• In patients with secondary hypogonadism (low testosterone with low/normal LH), measure: 4
  • TSH and free T4 (thyroid function)
  • Morning cortisol (adrenal function)
  • IGF-1 (growth hormone axis)
  • This distinguishes isolated gonadotropin deficiency from panhypopituitarism

Estradiol

• Measure serum estradiol only in testosterone-deficient patients who present with: 1, 2
  • Breast symptoms
  • Gynecomastia
• Perform this prior to starting testosterone therapy 1
• Elevated baseline estradiol requires endocrinology referral 1

Pre-Treatment Safety Laboratory Tests

Before initiating testosterone therapy, obtain: 2, 5

• Hemoglobin/hematocrit (baseline for polycythemia monitoring)
• Prostate-specific antigen (PSA) in men over 40 years
• Complete blood count (assess for erythrocytosis)
• Lipid panel (cardiovascular risk assessment)

Common Pitfalls to Avoid

• Do not diagnose hypogonadism based on a single testosterone measurement - biological variation requires confirmation 1, 2
• Do not measure testosterone in the afternoon or evening - levels decline throughout the day and may yield false-positive results 1
• Do not skip LH measurement - this is essential for determining etiology and guides whether additional pituitary evaluation is needed 1, 4
• Do not use screening questionnaires as a substitute for laboratory testing - they have poor specificity and sensitivity 1
• Do not forget to measure prolactin in secondary hypogonadism - missing a prolactinoma can have serious consequences 1, 4