Bioavailable Testosterone: Which Tests to Order Alongside It

What Bioavailable Testosterone Actually Measures

Bioavailable testosterone is the sum of free testosterone and albumin-bound testosterone. It excludes the roughly 44 to 65% of circulating testosterone that is tightly bound to SHBG and therefore unavailable to tissues. The Endocrine Society defines it as the "biologically active" fraction because albumin-bound testosterone dissociates readily in capillary beds and enters cells alongside the free fraction. Bhasin S, et al. Testosterone Therapy in Men with Hypogonadism: An Endocrine Society Clinical Practice Guideline. J Clin Endocrinol Metab. 2018.

Why Total Testosterone Misleads

Total testosterone includes SHBG-bound hormone that cannot act on androgen receptors. A man with obesity-related low SHBG may show a normal total testosterone of 400 ng/dL while his bioavailable fraction is frankly deficient. Conversely, a post-menopausal woman on oral estrogen may show an elevated SHBG that suppresses bioavailable testosterone well below 0.5 ng/dL even when her total testosterone reads in the normal range. Rosner W, et al. Position statement: Utility, limitations and pitfalls in measuring testosterone. J Clin Endocrinol Metab. 2007;92(2):405-413.

Calculated vs. Direct Assay

The Vermeulen equation calculates bioavailable testosterone from total testosterone, SHBG, and albumin (assumed 4.3 g/dL unless measured). Direct analog immunoassays for bioavailable testosterone suffer from poor precision at low concentrations and are not recommended for clinical use by the Endocrine Society. Vermeulen A, et al. A critical evaluation of simple methods for the estimation of free testosterone in serum. J Clin Endocrinol Metab. 1999;84(10):3666-3672.

The Core Paired Tests: What to Always Order Together

Total Testosterone (Serum)

Total testosterone is the arithmetic prerequisite for calculating bioavailable testosterone. The Endocrine Society guideline recommends measuring total testosterone by a reliable assay such as liquid chromatography-tandem mass spectrometry (LC-MS/MS), particularly when values fall below 400 ng/dL or when the clinical picture diverges from the number. Bhasin S, et al. J Clin Endocrinol Metab. 2018. Most commercial immunoassays overestimate total testosterone at concentrations below 150 ng/dL, a well-documented accuracy problem that compounds errors in the downstream bioavailable calculation.

Morning collection (7 to 10 AM) is standard because testosterone peaks early in the day. A single morning draw showing values below the lab's reference range should be confirmed with a second sample on a different day before clinical decisions are made.

Sex Hormone-Binding Globulin (SHBG)

SHBG is not optional. It is the primary driver of bioavailable testosterone variation independent of total testosterone production. Conditions that raise SHBG include hyperthyroidism, liver cirrhosis, aging, oral estrogen use, and anticonvulsant therapy. Conditions that lower SHBG include insulin resistance, obesity, hypothyroidism, nephrotic syndrome, and exogenous androgen use. Hammond GL. Diverse roles for sex hormone-binding globulin in reproduction. Biol Reprod. 2011;85(3):431-441.

The normal SHBG range in adult men is approximately 10 to 57 nmol/L and in premenopausal women approximately 18 to 144 nmol/L, though reference ranges vary by laboratory and assay platform.

Albumin

Albumin is either measured directly or assumed at 4.3 g/dL in the Vermeulen calculation. In patients with liver disease, nephrotic syndrome, malnutrition, or critical illness, measured albumin can fall below 3.0 g/dL, shifting the bioavailable testosterone result meaningfully. A serum albumin drawn alongside the testosterone panel costs under $5 and eliminates this source of calculation error. Vermeulen A, et al. J Clin Endocrinol Metab. 1999.

The Hypothalamic-Pituitary Axis Tests

Luteinizing Hormone (LH) and Follicle-Stimulating Hormone (FSH)

LH and FSH tell you where the problem originates. Low bioavailable testosterone with low or normal LH points to secondary (central) hypogonadism, involving the pituitary or hypothalamus. Low bioavailable testosterone with elevated LH points to primary hypogonadism, meaning testicular failure. This distinction changes treatment completely. The American Association of Clinical Endocrinologists (AACE) recommends measuring both gonadotropins in every patient with suspected hypogonadism before initiating androgen therapy. Petak SM, et al. AACE Medical Guidelines for Clinical Practice for the Evaluation and Treatment of Male Sexual Dysfunction. Endocr Pract. 2003;9(1):77-95.

FSH is the primary marker for spermatogenesis. Elevated FSH predicts impaired spermatogenesis and Sertoli cell dysfunction even when bioavailable testosterone is still within range. Men considering future fertility should have FSH documented before any androgen therapy is started.

Prolactin

A prolactinoma is a correctable cause of secondary hypogonadism that will not respond to exogenous testosterone. Serum prolactin should be measured once in any patient with newly identified secondary hypogonadism. A prolactin above 200 ng/mL in the absence of medication confounders (antipsychotics, metoclopramide, domperidone) warrants pituitary MRI. Melmed S, et al. Diagnosis and Treatment of Hyperprolactinemia: An Endocrine Society Clinical Practice Guideline. J Clin Endocrinol Metab. 2011;96(2):273-288.

Metabolic and Contextual Tests

Estradiol (E2)

Testosterone aromatizes to estradiol. In men on testosterone therapy, monitoring E2 is necessary because supraphysiologic estradiol causes gynecomastia, water retention, and mood instability. The AACE recommends measuring estradiol in men with symptoms of estrogen excess or deficiency. In women, estradiol contextualizes the androgen-to-estrogen ratio, which is relevant for PCOS assessment and menopause management. Seftel A. Male hypogonadism. Part I: Epidemiology of hypogonadism. Int J Impot Res. 2006;18(2):115-120.

Sensitive estradiol assays (LC-MS/MS or the "sensitive" Roche Elecsys method) are preferred in men because standard immunoassays have poor precision below 30 pg/mL.

Complete Metabolic Panel (CMP) and CBC

A complete metabolic panel screens for liver disease (which elevates SHBG and alters testosterone metabolism), kidney disease (which lowers albumin), and glucose abnormalities. Testosterone deficiency is associated with insulin resistance; the T2D Prevention Program data showed that men with low testosterone had a 42% higher risk of progressing to type 2 diabetes over 15 years. Selvin E, et al. Meta-analysis: glycosylated hemoglobin and cardiovascular disease in diabetes mellitus. Ann Intern Med. 2004;141(6):421-431.

A complete blood count (CBC) is essential before and during testosterone therapy because testosterone stimulates erythropoiesis. Hematocrit above 54% is a contraindication to continuing therapy per Endocrine Society guidance. Bhasin S, et al. J Clin Endocrinol Metab. 2018.

Thyroid-Stimulating Hormone (TSH)

Thyroid dysfunction directly alters SHBG. Hypothyroidism lowers SHBG, inflating the bioavailable testosterone fraction; hyperthyroidism raises SHBG, suppressing it. Undiagnosed thyroid disease can produce a false-positive low or high bioavailable testosterone result. The AACE recommends a TSH as part of the initial workup for unexplained fatigue and libido changes before attributing those symptoms to testosterone deficiency. Garber JR, et al. Clinical practice guidelines for hypothyroidism in adults. Endocr Pract. 2012;18(6):988-1028.

Fasting Glucose and HbA1c

Insulin resistance suppresses SHBG independently of testosterone production. A patient with an HbA1c of 6.2% and low SHBG may show a high bioavailable testosterone purely from SHBG suppression, not from true androgen excess. This distinction matters in women with PCOS, where the androgen-SHBG-insulin relationship drives most of the clinical phenotype. Diamanti-Kandarakis E, et al. Insulin resistance and the polycystic ovary syndrome revisited: An update on mechanisms and implications. Endocr Rev. 2012;33(6):981-1030.

Condition-Specific Add-On Tests

For Men Presenting with Possible Hypogonadism

A standard male androgen panel adds prostate-specific antigen (PSA) for men 40 and older. The Endocrine Society lists PSA above 4 ng/mL (or above 3 ng/mL in high-risk men) as a relative contraindication to testosterone initiation pending urology evaluation. Bhasin S, et al. J Clin Endocrinol Metab. 2018. Bone mineral density (DXA scan) is appropriate when bioavailable testosterone has been chronically low, given that androgen deficiency is a recognized contributor to osteoporosis in men. The International Society for Clinical Densitometry recommends DXA in men with hypogonadism regardless of age.

For Women with Suspected Androgen Excess (PCOS, Adrenal Pathology)

Women presenting with hirsutism, acne, or irregular cycles need DHEA-S and 17-hydroxyprogesterone added to the panel. DHEA-S separates adrenal from ovarian androgen sources. A DHEA-S above 700 mcg/dL suggests an adrenal tumor until proven otherwise. Morning 17-OHP above 2 ng/mL warrants an ACTH stimulation test to rule out non-classic congenital adrenal hyperplasia. The Endocrine Society PCOS guideline explicitly recommends both markers. Legro RS, et al. Diagnosis and Treatment of Polycystic Ovary Syndrome: An Endocrine Society Clinical Practice Guideline. J Clin Endocrinol Metab. 2013;98(12):4565-4592.

Androstenedione may also be added when ovarian androgen production is specifically in question, as it reflects both adrenal and ovarian secretion.

For Patients on TRT or Hormone Therapy

Monitoring panels after testosterone therapy initiation should include total testosterone, bioavailable testosterone, SHBG, estradiol, hematocrit, PSA (men), and lipid panel. The Endocrine Society recommends checking these at 3 and 6 months after initiation, then annually once stable. Ferritin is worth adding for men on injectable testosterone who show erythrocytosis, since iron-deficiency erythrocytosis has a different management pathway than testosterone-driven polycythemia. Bhasin S, et al. J Clin Endocrinol Metab. 2018.

Normal Bioavailable Testosterone Ranges

Reference ranges for bioavailable testosterone depend on assay method, age, and sex. The most cited reference values in the clinical literature are:

Men (Endocrine Society / Vermeulen-calculated):

• Ages 20 to 29: approximately 83 to 257 ng/dL
• Ages 40 to 49: approximately 60 to 220 ng/dL
• Ages 60 to 69: approximately 40 to 180 ng/dL

Women (premenopausal, Vermeulen-calculated):

• Follicular phase: approximately 0.6 to 3.8 ng/dL
• Postmenopausal (off HRT): approximately 0.2 to 1.8 ng/dL

The Endocrine Society 2018 guideline states: "We recommend against making a diagnosis of androgen deficiency in men who have total testosterone concentrations in the normal range without measurement of free or bioavailable testosterone in those with conditions that alter SHBG concentrations." Bhasin S, et al. J Clin Endocrinol Metab. 2018.

Age-matched ranges matter because bioavailable testosterone declines roughly 2 to 3% per year after age 30, even when total testosterone appears stable, owing largely to the age-related rise in SHBG. Harman SM, et al. Longitudinal effects of aging on serum total and free testosterone levels in healthy men. J Clin Endocrinol Metab. 2001;86(2):724-731.

How to Raise Low Bioavailable Testosterone

Low bioavailable testosterone has two correctable levers: increase total testosterone production or decrease SHBG. Interventions that address both are clinically preferred.

Lifestyle Modifications

Resistance training three to five sessions per week is associated with a 15 to 20% increase in bioavailable testosterone over 12 weeks in sedentary middle-aged men, mediated partly through SHBG reduction. Kraemer WJ, et al. Hormonal and growth factor responses to heavy resistance exercise protocols. J Appl Physiol. 1990;69(4):1442-1450. Fat loss also raises bioavailable testosterone by lowering SHBG through improved insulin sensitivity. Sleep optimization (7 to 9 hours) supports the nocturnal testosterone pulse that drives morning peak values.

Zinc deficiency is the one micronutrient deficiency with solid evidence for depressing testosterone. Prasad AS, et al. Zinc status and serum testosterone levels of healthy adults. Nutrition. 1996;12(5):344-348. Men with documented deficiency who supplement zinc at 25 to 45 mg/day may see partial recovery of testicular testosterone output within 8 to 12 weeks.

Pharmacological Options

When bioavailable testosterone remains low after optimizing modifiable factors, testosterone replacement therapy is the standard of care. In men with confirmed hypogonadism (bioavailable testosterone below age-adjusted reference ranges on two morning draws), the Endocrine Society recommends testosterone therapy to achieve serum total testosterone in the mid-normal range (400 to 700 ng/dL). Available formulations include testosterone cypionate or enanthate (50 to 100 mg IM weekly or 75 to 200 mg every two weeks), testosterone undecanoate 750 mg IM every 10 weeks, transdermal 1.62% gel (20.25 to 81 mg/day), and transdermal patches (2 to 6 mg/day). Bhasin S, et al. J Clin Endocrinol Metab. 2018.

Clomiphene citrate 25 to 50 mg every other day or daily raises LH-driven endogenous testosterone and may be preferred in men who want to preserve fertility. It is off-label for this indication but widely used. Enclomiphene, the active trans-isomer, has shown total testosterone increases of approximately 140 ng/dL over placebo in phase 2 trials. Kim ED, et al. A randomized, double-blind, placebo-controlled phase II trial of clomiphene citrate for the treatment of idiopathic hypogonadism. Urology. 2013;82(6):1251-1256.

How to Lower Elevated Bioavailable Testosterone

Elevated bioavailable testosterone in women commonly reflects PCOS-related ovarian androgen excess or low SHBG from insulin resistance, not a production problem alone. Treatment targets the underlying driver.

Raising SHBG

Oral combined contraceptives containing ethinyl estradiol raise SHBG by two- to fourfold, reducing bioavailable testosterone substantially. This is the pharmacologic basis for using combined OCs to treat hirsutism in PCOS. Falsetti L, et al. Long-term treatment with a low-dose oral contraceptive in polycystic ovary syndrome. Gynecol Endocrinol. 1995;9(3):189-196. Spironolactone 50 to 200 mg/day blocks androgen receptors and mildly raises SHBG, providing a dual mechanism. The Endocrine Society PCOS guideline recommends combined OCs as first-line pharmacotherapy for hyperandrogenism. Legro RS, et al. J Clin Endocrinol Metab. 2013.

Addressing Insulin Resistance

Metformin 500 to 2,000 mg/day lowers insulin levels, which raises SHBG and reduces ovarian androgen production in women with PCOS. A Cochrane review of 41 randomized trials found metformin produced statistically significant reductions in fasting insulin, free androgen index, and LH/FSH ratio compared with placebo. Costello MF, et al. A systematic review and meta-analysis of randomized controlled trials on metformin co-administration during gonadotrophin ovulation induction. Hum Reprod. 2006;21(6):1387-1399. Weight loss of even 5 to 10% of body weight consistently raises SHBG and lowers bioavailable testosterone in overweight women with PCOS.

The Complete Recommended Panel at a Glance

The following panel reflects the minimum markers for a thorough androgen assessment, consistent with Endocrine Society and AACE guidance:

| Test | Purpose | Required For | |---|---|---| | Total testosterone (AM) | Calculation input | All patients | | SHBG | Calculation input | All patients | | Serum albumin | Calculation refinement | Liver/renal disease | | LH | Axis localization | All new presentations | | FSH | Axis localization, fertility | All new presentations | | Prolactin | Rule out prolactinoma | Secondary hypogonadism | | Estradiol (sensitive assay) | Aromatization monitoring | Men on TRT; women | | TSH | SHBG confounder | Fatigue/libido symptoms | | HbA1c / fasting glucose | SHBG confounder, metabolic | All patients | | CBC | Erythrocytosis monitoring | TRT initiation/follow-up | | CMP | Liver/kidney, albumin | All new presentations | | PSA | Prostate safety | Men 40+ before TRT | | DHEA-S | Adrenal vs. Ovarian source | Women with androgen excess | | 17-OHP (AM) | Rule out NCAH | Women with androgen excess |

Drawing all applicable tests from one morning fasting blood draw minimizes variability. Total testosterone and SHBG fluctuate with diurnal rhythms and recent meals; a fasting state stabilizes albumin and glucose-related SHBG suppression for cleaner Vermeulen calculations.