Free Testosterone Measurement
Equilibrium Dialysis vs Vermeulen Calculation vs Direct Analog Immunoassay
Evidence grade 4 of 5, Strong. Read how the Atlas grades evidence.
The written rationale for this grade is still being prepared.
- FDA
- Methods run in CLIA labs; equilibrium dialysis is the reference standard for free T
Reference only. Not medical advice.
What can go wrong
A 'low free testosterone' produced by a direct analog immunoassay may not be reproducible by equilibrium dialysis or calculation and should not by itself justify therapy. Calculated free T also depends on the accuracy of the underlying total-T and SHBG assays; garbage in, garbage out. Free-T thresholds are less standardized than total-T thresholds, which clinics can exploit.
This is something to take to a clinician. The Atlas can tell you what is known about free testosterone measurement in general. It cannot tell you what is happening in your body, and the difference matters most for exactly the symptoms that bring people to this page.
Free testosterone (the non-protein-bound fraction) is measured three ways with very different reliability. Equilibrium dialysis (with mass-spec quantitation) is the gold standard. A calculated free testosterone from total T, SHBG, and albumin - most commonly the Vermeulen 1999 equation - is a validated, widely recommended practical alternative. The discredited method is the direct analog (single-step) immunoassay, which the Endocrine Society explicitly advises against because it correlates poorly with reference measurements. Because optimization clinics often choose the cheapest direct method or a favorable calculator, 'low free T' results can vary wildly for the same blood.
The reason two clinics can call the same man 'low free T' and 'normal.'
What people take it for
What it is taken for. Approval, where it exists, covers only some of these.
How it works
Equilibrium dialysis physically separates free from bound hormone across a membrane, then quantifies the free fraction. The Vermeulen calculation estimates free T from total T and binding-protein concentrations using known SHBG/albumin association constants. The direct analog immunoassay uses a labeled testosterone analog meant to track only the free fraction, but the analog's binding behavior is unreliable, producing values that diverge from reference methods.
Same blood, three answers
Because SHBG drives the free fraction, a man with high or low SHBG can look normal on total T but be flagged 'low free T' on a direct analog assay that a clinic chose for cost. The Endocrine Society 2018 guideline recommends free testosterone by equilibrium dialysis or by a reliable calculated estimate, and specifically discourages the direct analog method. A clinic that reports calculated free T should also disclose which calculator and which SHBG/albumin assay it used, since those propagate into the number that decides therapy.
What people get wrong
Often said'Low free T' is a precise, settled measurement
ActuallyIt is method-dependent and threshold-soft. The same sample can read normal by dialysis or calculation and low by a discredited analog assay.
Often saidFree T is the number that really matters, so trust whatever the panel reports
ActuallyFree T is informative when SHBG is abnormal, but only if measured by a reliable method; a direct analog result is not a trustworthy basis for diagnosis.
Who profits, and who pushes back
Alex Vermeulen, MD
The calculated value tracks equilibrium dialysis closely and is preferable to flawed direct immunoassays.
Endocrine Society
Free testosterone should be estimated by equilibrium dialysis or accurate calculation, not by direct analog immunoassay.
What we do not know
- How widely do consumer panels still report direct analog free testosterone?
- How much do calculated free-T values shift between competing online calculators using different constants?
At a glance
- Gold standard
- Equilibrium dialysis + LC-MS/MS
- Recommended practical method
- Calculated free T (Vermeulen 1999)
- Discredited method
- Direct analog/single-step immunoassay
- Inputs for calculation
- Total T, SHBG, albumin
What this entry does not yet cover
What follows is unfinished work on our side, not a finding about Free Testosterone Measurement. A short entry is not a safe entry.
- Timeline. We have not built the dated record of research and regulatory events.
- Cost and market. We have not documented what it sells for or who sells it.
- Supply chain. We have not mapped where it is made.
- Who uses it. We have not documented the user groups.
- What could change. No forward-looking summary yet.
- How it compares. We have not set it against the alternatives.
- Children and teenagers. We have not checked what is known about younger people.
Sources
The studies below are named and described, but not linked. You cannot open them from here and check our reading against them, which means this page is our summary of the literature rather than a document you can verify. Weigh it accordingly until the links exist.
The studies that matter
Vermeulen, Verdonck, Kaufman1999
Published the widely used calculated free testosterone equation, validated against equilibrium dialysis
Endocrine Society guideline (free T measurement)2018
Recommends equilibrium dialysis or accurate calculation; advises against the direct analog immunoassay
Rosner et al. assay position statement2007
Criticized direct/analog free-testosterone immunoassays as inaccurate