Is the calculated free testosterone (FT) level still accurate in patients with high Sex Hormone-Binding Globulin (SHBG) levels?

Accuracy of Free Testosterone Calculation in Patients with High SHBG

Calculated free testosterone (FT) remains accurate in patients with high Sex Hormone-Binding Globulin (SHBG) levels, with calculated FT showing high diagnostic accuracy (sensitivity 89%, specificity 83%) for detecting hyperandrogenism even in conditions with altered SHBG levels. 1

Understanding Free Testosterone and SHBG

Free testosterone represents only 1-2% of total testosterone in circulation but is the biologically active form responsible for androgen action 2. The majority of testosterone in plasma is bound to proteins:

• Approximately 40% bound to SHBG
• About 58% loosely bound to albumin
• Only 2% remains unbound (free) 3

SHBG levels can be altered in various conditions:

• Increased SHBG: Liver disease, hyperthyroidism, pregnancy, estrogen therapy
• Decreased SHBG: Obesity, insulin resistance, hypothyroidism, androgen therapy

Evidence Supporting Accuracy of Calculated Free Testosterone

The 2025 Human Reproduction Update guidelines provide strong evidence that calculated free testosterone (cFT) is a reliable diagnostic marker for hyperandrogenism with high accuracy:

• Sensitivity: 0.89 (95% CI: 0.74-0.98)
• Specificity: 0.83 (95% CI: 0.77-0.87)
• AUC: 0.85 (95% CI: 0.81-0.88) 1

This diagnostic accuracy was maintained across different SHBG levels, making cFT a reliable marker even in conditions with altered binding proteins.

The original Vermeulen equation for calculating free testosterone has been validated against equilibrium dialysis (the gold standard method) and shows excellent correlation 4. This equation accounts for SHBG concentration in its calculation, making it reliable even when SHBG levels are abnormal.

Methods for Measuring/Calculating Free Testosterone

1. Equilibrium Dialysis (Gold Standard):

   • Direct measurement of free testosterone
   • Labor-intensive, costly, and not widely available
   • Most accurate method 5

2. Calculated Free Testosterone (cFT):

   • Uses total testosterone and SHBG measurements
   • Employs equations (e.g., Vermeulen) to calculate free testosterone
   • Highly correlated with equilibrium dialysis results 4
   • Widely available and practical for clinical use

3. Free Androgen Index (FAI):

   • Simple ratio of total testosterone to SHBG (100 × T/SHBG)
   • Less accurate than cFT, especially with varying SHBG levels 4
   • Still clinically useful with pooled sensitivity of 0.78 and specificity of 0.85 1, 6

4. Direct Free Testosterone Immunoassays:

   • Less reliable and not recommended
   • Values are only a fraction of true free testosterone 4

Clinical Considerations

When interpreting calculated free testosterone in patients with high SHBG:

• Assay Method Matters: LC-MS/MS measurement of total testosterone provides superior accuracy compared to immunoassays 1
• Calculation Method: The Vermeulen equation is most validated for calculating free testosterone 4
• Special Circumstances: During pregnancy, calculated FT may underestimate true free testosterone due to estradiol occupying SHBG binding sites 4

Algorithm for Testosterone Assessment in High SHBG States

1. Measure total testosterone using LC-MS/MS (preferred) or high-quality immunoassay
2. Measure SHBG using a reliable immunoassay
3. Calculate free testosterone using the Vermeulen equation
4. Interpret results in clinical context, considering that:
   • cFT is reliable even with high SHBG
   • If very high SHBG (>120 nmol/L) or pregnancy, consider direct measurement by equilibrium dialysis if available

Common Pitfalls to Avoid

• Relying solely on total testosterone: In conditions with altered SHBG, total testosterone may not reflect bioavailable hormone
• Using direct free testosterone immunoassays: These are inaccurate and provide values that are only a fraction of true free testosterone 4
• Using FAI alone: While clinically useful, FAI is less accurate than calculated free testosterone, especially with varying SHBG levels 4
• Ignoring assay method: Immunoassays for total testosterone are less accurate than LC-MS/MS, particularly at lower concentrations seen in women 1

In conclusion, calculated free testosterone remains a reliable and accurate method for assessing bioavailable testosterone even in patients with high SHBG levels, provided that high-quality assays are used for measuring total testosterone and SHBG.