What are the best practices when testing for testosterone levels?

From the Guidelines

The best practice for testing testosterone involves measuring total testosterone levels in the morning, using a cut-off of 300 ng/dL, and confirming the diagnosis with two separate morning measurements, as recommended by the American Urological Association 1. This approach ensures accurate diagnosis and appropriate treatment planning. The diagnosis of low testosterone should be made only after two total testosterone measurements are taken on separate occasions, with both conducted in an early morning fashion, and in the presence of symptoms and/or signs 1. Key considerations include:

• Measuring total testosterone levels in the morning, when levels are naturally highest, preferably after fasting
• Using a cut-off of 300 ng/dL to support the diagnosis of low testosterone
• Confirming the diagnosis with two separate morning measurements, ideally on different days
• Considering additional tests, such as luteinizing hormone (LH), follicle-stimulating hormone (FSH), prolactin, and sometimes a complete metabolic panel, to help determine the cause of low testosterone
• Using the most reliable testing methods, such as liquid chromatography-mass spectrometry (LC-MS/MS) or high-quality immunoassays The most recent and highest quality study, from 2024, supports these recommendations and highlights the importance of accurate and reliable testing methods 1. It is essential to note that the diagnosis of testosterone deficiency requires both a low testosterone measurement and the presence of select symptoms and/or signs, as outlined in the American Urological Association guidelines 1. Additionally, clinicians should consider measuring total testosterone in patients with a history of unexplained anemia, bone density loss, diabetes, exposure to chemotherapy, exposure to testicular radiation, HIV/AIDS, chronic narcotic use, male infertility, pituitary dysfunction, and chronic corticosteroid use, even in the absence of symptoms or signs associated with testosterone deficiency 1.

From the FDA Drug Label

DOSAGE AND ADMINISTRATION Prior to initiating testosterone enanthate injection, confirm the diagnosis of hypogonadism by ensuring that serum testosterone concentrations have been measured in the morning on at least two separate days and that these serum testosterone concentrations are below the normal range The best practice when testing testosterone is to measure serum testosterone concentrations in the morning on at least two separate days to confirm the diagnosis of hypogonadism, as stated in the dosage and administration section of the drug label 2.

• Key points:
  • Measure serum testosterone concentrations
  • Measure in the morning
  • Measure on at least two separate days
  • Ensure concentrations are below the normal range to confirm hypogonadism

From the Research

Best Practice for Testing Testosterone

• Total testosterone should be measured in a fasting state in the morning using a reliable assay, such as liquid chromatography tandem mass spectrometry, in a laboratory that is certified by an accuracy-based benchmark 3.
• Free testosterone levels should be measured in men in whom alterations in binding protein concentrations are suspected or in whom total testosterone levels are only slightly above or slightly below the lower limit of the normal male range for testosterone 3.
• The circulating concentrations of total and free testosterone vary substantially in people over time due to biologic factors as well as due to measurement variation, making accurate measurement essential for diagnosis of androgen disorders 3.

Considerations for Hypogonadal Patients

• Testosterone replacement therapy can increase the risk of infertility, but human chorionic gonadotropin (HCG) therapy may be a viable option for management of secondary hypogonadism and male infertility 4.
• HCG therapy can trigger testosterone and sperm production without the side effects on fertility seen in testosterone replacement therapy 4.
• Concomitant intramuscular HCG can preserve spermatogenesis in men undergoing testosterone replacement therapy 5.
• Alternatives to testosterone replacement therapy, such as clomiphene citrate and HCG, have been shown to increase endogenous testosterone production and may be considered for young hypogonadal patients who desire future fertility 6.