Testosterone Testing in Men: When to Check, How to Interpret, and the Prescribing Minefield of TRT

 

Testosterone Testing in Men: When to Check, How to Interpret, and the Prescribing Minefield of TRT

A total testosterone of 280 at 8 AM in a symptomatic man means something. A total testosterone of 280 drawn at 3 PM after a night of poor sleep means almost nothing.

Testosterone replacement therapy (TRT) is one of the fastest-growing prescriptions in men's health, driven by direct-to-consumer marketing, telehealth "optimization" clinics, and patient demand. For primary care NPs, this creates a two-sided problem: underdiagnosis of true hypogonadism in men who would benefit from treatment, and overprescription of testosterone in men who don't meet diagnostic criteria and face real risks. Getting the workup right is the foundation of safe prescribing.

The Physiology: What Testosterone Does and How It's Regulated

The hypothalamic-pituitary-gonadal (HPG) axis works as a feedback loop: GnRH from the hypothalamus stimulates LH and FSH from the anterior pituitary, LH stimulates Leydig cells in the testes to produce testosterone, and testosterone feeds back to suppress GnRH and LH. Understanding this loop is essential because it determines whether hypogonadism is primary (testicular failure, high LH) or secondary (pituitary/hypothalamic dysfunction, low/normal LH)—and the distinction changes the workup and treatment approach entirely.

Testosterone circulates in three forms: tightly bound to sex hormone-binding globulin (SHBG, ~44%), loosely bound to albumin (~54%), and free/unbound (~2–3%). Free testosterone plus albumin-bound testosterone together constitute "bioavailable testosterone"—the fraction that is biologically active. Total testosterone measures all three forms. In conditions that alter SHBG (and there are many), total testosterone can be misleading.

When to Test: The Appropriate Indications

Test for Hypogonadism When

• Specific symptoms are present: Decreased libido, erectile dysfunction, reduced morning erections, fatigue, decreased energy, loss of muscle mass/strength, increased body fat (especially visceral), depressed mood, difficulty concentrating, gynecomastia, decreased bone mineral density/fragility fractures, infertility, decreased testicular volume
• Conditions associated with hypogonadism: Type 2 diabetes (prevalence of hypogonadism ~25–40%), obesity (BMI >30, especially >40), metabolic syndrome, HIV/AIDS, chronic opioid use, glucocorticoid therapy, pituitary disease/mass, Klinefelter syndrome (47,XXY), prior testicular injury/orchitis/torsion, prior chemotherapy or radiation, hemochromatosis, obstructive sleep apnea

Do NOT screen asymptomatic men. The AUA, Endocrine Society, and ACP all recommend against population-based testosterone screening. Testing should be symptom-driven.

How to Test: Getting It Right the First Time

Step 1: Total Testosterone (Morning Draw)

The Timing Rule

Testosterone follows a circadian rhythm, peaking between 6–10 AM and declining 20–25% by afternoon. A total testosterone drawn at 3 PM will be significantly lower than the same man's 8 AM level. Always draw between 6–10 AM, fasting, after a normal night of sleep. Acute illness, sleep deprivation, recent heavy exercise, and alcohol use within 24 hours all suppress testosterone transiently and should prompt rescheduling. In men over 40, the circadian variation blunts somewhat, but morning draw remains standard practice.

• Test: Total testosterone by liquid chromatography-tandem mass spectrometry (LC-MS/MS). This is the gold standard. Immunoassay methods are less accurate, especially at low levels. If your lab uses immunoassay, be aware of potential inaccuracy and consider sending to a reference lab.
• Confirm with a second sample: A single low total testosterone is not diagnostic. The Endocrine Society requires two morning total testosterone levels on separate days before diagnosing hypogonadism. Up to 30% of men with an initially low level will have a normal level on repeat testing.

Step 2: Classify the Level

Total Testosterone	Interpretation	Next Step
>350 ng/dL	Normal for most assays and guidelines	Testosterone deficiency is unlikely as the cause of symptoms. Investigate other etiologies (depression, sleep apnea, thyroid, anemia).
250–350 ng/dL	Equivocal / "gray zone"	Check free testosterone (or calculate it) AND SHBG. If free T is low, hypogonadism is supported. If free T is normal, the low total T is due to low SHBG, not true deficiency.
<250 ng/dL	Unequivocally low	Confirm on a second morning draw. If confirmed, proceed with classification (primary vs. secondary) and evaluate for underlying causes before prescribing.

The SHBG Confounders

SHBG binds testosterone tightly, making it biologically unavailable. Conditions that raise SHBG inflate total testosterone (making the man appear replete when he isn't), and conditions that lower SHBG deflate total testosterone (making the man appear deficient when free T is actually normal).

• SHBG increases (total T falsely reassuring): aging, liver disease/cirrhosis, hyperthyroidism, anticonvulsants (phenytoin, carbamazepine), HIV, estrogen therapy, anorexia
• SHBG decreases (total T falsely low): obesity, type 2 diabetes, metabolic syndrome, hypothyroidism, nephrotic syndrome, exogenous androgens, glucocorticoids, GH excess

In any man with obesity, diabetes, or a total T in the 250–350 gray zone, you must check free testosterone or SHBG to interpret the total T accurately. Calculated free testosterone (using the Vermeulen equation with total T, SHBG, and albumin) is preferred over direct free T immunoassay, which is notoriously inaccurate.

Step 3: Primary vs. Secondary Hypogonadism

Type	FSH/LH	Cause	Key Workup
Primary (Hypergonadotropic)	Elevated LH and FSH (the pituitary is screaming at the testes and they're not responding)	Klinefelter syndrome (47,XXY), prior orchitis (mumps), testicular torsion/trauma, chemotherapy/radiation, cryptorchidism, age-related (late-onset hypogonadism)	Karyotype if <40 with primary hypogonadism (Klinefelter is the most common genetic cause, ~1 in 600 men, and the majority are undiagnosed). Testicular exam. Semen analysis if fertility desired.
Secondary (Hypogonadotropic)	Low or inappropriately normal LH and FSH	Pituitary adenoma (prolactinoma most common), hemochromatosis, Kallmann syndrome, obesity, opioid use, glucocorticoids, chronic illness, idiopathic	Prolactin (elevated = pituitary adenoma until proven otherwise). Iron/ferritin (hemochromatosis is a treatable cause). MRI of the pituitary if prolactin elevated, very low testosterone (<150), or signs of mass effect (visual field changes, headache). Evaluate for other pituitary hormone deficiencies (cortisol, TSH).

Never Skip the Classification

Prescribing TRT without checking LH/FSH is a missed opportunity to diagnose a prolactinoma, hemochromatosis, or Klinefelter syndrome—all of which have specific treatments beyond testosterone. A prolactinoma causing secondary hypogonadism may be cured with cabergoline. Hemochromatosis causing secondary hypogonadism is treated with phlebotomy, and testosterone may recover. LH, FSH, and prolactin are mandatory before prescribing TRT.

The Complete Pre-TRT Workup

Test	Why
Total testosterone (AM, ×2)	Confirm deficiency on two separate morning draws
Free testosterone (calculated) or SHBG	Clarify equivocal total T; essential in obesity, diabetes, liver disease
LH, FSH	Classify primary vs. secondary. Identify treatable pituitary/hypothalamic causes.
Prolactin	Screen for prolactinoma (most common secretory pituitary adenoma in men)
CBC (hemoglobin/hematocrit)	Baseline before TRT (testosterone stimulates erythropoiesis; polycythemia is the most common adverse effect). Hematocrit >50% is a relative contraindication to starting TRT.
PSA	Baseline prostate cancer screening. TRT is contraindicated in known/suspected prostate cancer. PSA >4 (or >3 in high-risk men) warrants urology referral before starting TRT.
Lipid panel	Baseline cardiovascular risk assessment
BMP	Baseline renal function
DEXA scan	If osteoporosis or fragility fracture is the presenting symptom
Iron/ferritin	Screen for hemochromatosis if secondary hypogonadism
Semen analysis	If fertility is desired. TRT suppresses spermatogenesis and is NOT appropriate for men who want to conceive.
TSH	Hypothyroidism mimics hypogonadal symptoms and raises SHBG

Testosterone Replacement: Formulations and Monitoring

Formulation	Route	Typical Dosing	Key Considerations
Testosterone cypionate	IM injection	100–200 mg every 1–2 weeks	Most common, least expensive. Peak-trough fluctuations (mood/energy swings). Self-injection taught to patients. Draw trough levels (just before next injection).
Testosterone enanthate	IM injection	100–200 mg every 1–2 weeks	Similar pharmacokinetics to cypionate. Interchangeable in practice.
Testosterone undecanoate (Aveed)	IM injection	750 mg initially, at 4 weeks, then every 10 weeks	Longer intervals. Requires REMS program (risk of pulmonary oil microembolism, anaphylaxis). Must be administered in clinic with 30-min observation.
Topical gel (AndroGel, Testim, Vogelxo)	Transdermal	1–4 pumps daily (varies by product)	Steady-state levels. Risk of transference to women/children through skin contact (FDA black box warning). Apply to shoulders/upper arms, wash hands, cover with clothing. Draw levels 2–4 hours after application.
Testosterone patch (Androderm)	Transdermal	2–4 mg nightly	Skin irritation common (~30%). Apply to back, abdomen, thigh, or upper arm. Rotate sites.
Nasal gel (Natesto)	Intranasal	5.5 mg per nostril TID	Short-acting. May preserve spermatogenesis better than other formulations (less HPG suppression due to pulsatile delivery). Nasal irritation.
Subcutaneous pellets (Testopel)	Implant	150–450 mg every 3–6 months	Inserted in office. Steady levels. Risk of pellet extrusion, infection at insertion site. Cannot adjust dose easily once implanted.
Oral testosterone undecanoate (Jatenzo)	Oral	158–396 mg BID with food	Newer formulation. Absorbed via lymphatic system (avoids first-pass hepatotoxicity of older oral forms). BP monitoring required (increases systolic ~3–5 mmHg).

Monitoring on TRT

Test	When	Target / Action
Total testosterone	3–6 months after initiation or dose change, then annually	Goal: mid-normal range (400–700 ng/dL). For injections, draw trough level. For gels, draw 2–4 hours post-application.
Hematocrit	3–6 months, then every 6–12 months	Hematocrit >54% = hold TRT, reduce dose, or phlebotomize. Polycythemia is the most common adverse effect and increases thrombotic risk (stroke, PE, DVT).
PSA	3–6 months, 12 months, then per screening guidelines	PSA rise >1.4 ng/mL within 12 months of starting TRT = urology referral. TRT does not cause prostate cancer, but may accelerate growth of occult cancer.
Lipid panel	6–12 months	TRT may lower HDL. Monitor cardiovascular risk profile.
Estradiol	If gynecomastia, nipple tenderness, or mood symptoms develop	Testosterone aromatizes to estradiol. Elevated estradiol (>40–50 pg/mL) may require dose reduction. Routine AI (anastrozole) use is NOT recommended by guidelines.
BMP	Annually	Renal function monitoring
Bone density (DEXA)	1–2 years if osteoporosis was the indication	Expect improvement in BMD within 12–24 months
Symptom assessment	Every visit	If no symptom improvement at 3–6 months with adequate levels, reconsider the diagnosis. Low testosterone may not have been the cause.

Contraindications to TRT

Do Not Prescribe TRT If

• Desire for fertility: Exogenous testosterone suppresses LH/FSH, which suppresses spermatogenesis. TRT is a form of male contraception. For hypogonadal men who want to conceive, use clomiphene citrate (off-label, stimulates endogenous testosterone production via HPG axis) or hCG (mimics LH, stimulates testicular testosterone production without suppressing spermatogenesis).
• Known or suspected prostate cancer (absolute)
• Breast cancer in men (absolute)
• Hematocrit >50% at baseline (relative; evaluate for other causes of polycythemia first)
• Untreated severe obstructive sleep apnea: TRT can worsen OSA. Treat OSA first, then reassess symptoms.
• Uncontrolled heart failure (NYHA Class III–IV): Risk of fluid retention
• PSA >4 ng/mL (or >3 in high-risk men) without urological evaluation
• Recent (within 3–6 months) major cardiovascular event: MI, stroke, DVT, PE. The cardiovascular safety of TRT is addressed by the TRAVERSE trial (see below).

The TRAVERSE Trial: Cardiovascular Safety Resolved

Practice-Changing Evidence

The TRAVERSE trial (2023, NEJM) was the first large, randomized, placebo-controlled cardiovascular outcomes trial of TRT. In 5,246 men aged 45–80 with hypogonadism and established or high-risk cardiovascular disease, testosterone gel did not increase the incidence of major adverse cardiovascular events (MACE: CV death, MI, stroke) compared to placebo over a median follow-up of 33 months. This resolved a decade of uncertainty following the 2010 TOM trial and observational studies that raised cardiovascular concerns. The 2024 updated AUA and Endocrine Society guidelines reflect this data: TRT is not contraindicated in men with stable cardiovascular disease who meet diagnostic criteria for hypogonadism. However, the trial did confirm increased rates of atrial fibrillation, AKI, and pulmonary embolism in the TRT group, so monitoring remains essential.

The Opioid-Induced Hypogonadism Problem

Chronic opioid use suppresses GnRH pulsatility, causing secondary hypogonadism in an estimated 50–90% of men on long-term opioids. This is one of the most underrecognized causes of testosterone deficiency in primary care.

• Mechanism: Opioids suppress the HPG axis centrally. LH and FSH are low. Testosterone is low.
• Symptoms: Sexual dysfunction, fatigue, depression, osteoporosis—often attributed to the underlying pain condition rather than the opioid.
• Management: First step is opioid dose reduction or discontinuation (with appropriate tapering). If opioids cannot be reduced, TRT may be considered, but it treats the symptom, not the cause. Alternatives: clomiphene (restores endogenous production in some patients).
• Screen all men on chronic opioid therapy (≥3 months) with a morning total testosterone if they report sexual dysfunction, fatigue, or mood changes.

Obesity and Testosterone: The Chicken-and-Egg Problem

Functional Hypogonadism

Obesity is the most common cause of low total testosterone in men. Excess adipose tissue aromatizes testosterone to estradiol (lowering T), and obesity lowers SHBG (making total T appear lower while free T may be preserved). This creates a functional or "reversible" hypogonadism. Weight loss of 5–10% can increase total testosterone by 50–100 ng/dL without TRT. Bariatric surgery patients often normalize testosterone. Before prescribing TRT to an obese man with a borderline-low total T, check free testosterone and counsel on weight loss. If free T is normal, the low total T is an SHBG effect, not true deficiency. The AUA 2024 guidelines recommend lifestyle intervention as first-line for obese men with functional hypogonadism.

Testosterone in Older Men: The Age-Related Decline Debate

Testosterone declines approximately 1–2% per year after age 30. By age 70, many men have total testosterone below the young-adult reference range. Whether this constitutes "disease" requiring treatment or a normal physiologic process remains debated. Key points:

• The Endocrine Society recommends against routine testosterone testing in men >65 based on age alone
• Test only if specific hypogonadal symptoms are present
• The TTrials (Testosterone Trials, 2016) showed modest benefits in sexual function and a small improvement in walking distance in men ≥65 with low T, but no meaningful improvement in vitality or cognitive function
• The TRAVERSE trial established cardiovascular safety but also showed increased AF, AKI, and PE risk
• In older men, the benefit-risk calculus is narrower. Start with the lowest effective dose and monitor hematocrit closely (polycythemia risk is highest in older men).

Clomiphene Citrate: The Fertility-Preserving Alternative

Clomiphene citrate (off-label in men) is a selective estrogen receptor modulator (SERM) that blocks estrogen feedback at the hypothalamus and pituitary, increasing GnRH, LH, and FSH secretion, which in turn stimulates endogenous testosterone production and preserves spermatogenesis.

• Indication: Secondary hypogonadism in men who desire fertility, or as a trial before committing to TRT
• Typical dose: 25–50 mg daily or every other day
• Monitoring: Total testosterone, LH, estradiol at 4–6 weeks. Adjust dose to achieve mid-normal testosterone without excessive estradiol.
• Advantages: Oral, inexpensive, preserves spermatogenesis, reversible
• Limitations: Not FDA-approved for male hypogonadism (off-label). May not be as effective as TRT for symptom relief. Elevated estradiol can cause gynecomastia. Contraindicated in primary hypogonadism (LH is already high—stimulating it further won't help).

The "Testosterone Clinic" Phenomenon and Scope of Practice

What You're Seeing in Practice

Patients are arriving at primary care already on TRT prescribed by telehealth or "men's health optimization" clinics, often without proper diagnostic workup. Common problems you'll encounter:

• Started on TRT without two confirmed low morning testosterone levels
• No LH/FSH checked (so primary vs. secondary was never classified, and treatable causes like prolactinoma were never evaluated)
• No baseline PSA or hematocrit
• Concurrent anastrozole prescribed routinely (not guideline-recommended; lowers estradiol and may worsen bone density)
• Concurrent hCG prescribed to "maintain fertility"—partially effective but adds cost and complexity
• Supraphysiologic dosing targeting the "high-normal" range or above
• No monitoring of hematocrit (polycythemia developing silently)

Your role: Don't reflexively discontinue their TRT, but do complete the workup that was skipped. Check LH/FSH (will be suppressed on TRT, but establish the record), PSA, hematocrit, and estradiol. Verify the indication was legitimate. Adjust to guideline-concordant dosing and monitoring.

The Pitfalls

• Drawing testosterone in the afternoon: The #1 collection error. An afternoon level can be 20–25% lower than the morning peak. Always 6–10 AM, fasting.
• Using a single low level to diagnose: The Endocrine Society requires two confirmed low morning levels. Up to 30% of men normalize on repeat testing.
• Ignoring SHBG in obese/diabetic men: Total testosterone is artificially low when SHBG is low. Check free testosterone before diagnosing.
• Skipping LH/FSH: This misses prolactinoma, hemochromatosis, and Klinefelter syndrome—all of which have specific treatments.
• Prescribing TRT to a man who wants children: TRT is a contraceptive. It suppresses spermatogenesis. Recovery after discontinuation is unpredictable (months to >1 year; some men have permanent impairment). Use clomiphene or hCG instead.
• Not monitoring hematocrit: Polycythemia is the most common adverse effect. Hematocrit >54% requires intervention (dose reduction, therapeutic phlebotomy, or discontinuation).
• Treating "low-normal" testosterone in the absence of symptoms: A total testosterone of 350 in a 60-year-old man who feels well, has normal libido, and has no metabolic risk factors does not require treatment. Treat the patient, not the number.
• Routine aromatase inhibitor use: Anastrozole to "manage estrogen" is not recommended by the Endocrine Society or AUA guidelines outside of specific clinical scenarios. It lowers estradiol, which is important for bone health and cardiovascular protection.
• Forgetting the transference risk: Topical testosterone gel can transfer to women and children through skin contact, causing virilization. FDA black box warning. Counsel on application, hand washing, covering the site, and laundering clothing/bedding.
• Missing the opioid connection: Chronic opioid use causes secondary hypogonadism in the majority of men. Check testosterone before attributing sexual dysfunction to "age" in a patient on chronic opioids.

NP Prescribing Considerations

Testosterone is a Schedule III controlled substance (DEA). NP prescriptive authority for controlled substances varies by state. Before prescribing:

• Verify your state scope-of-practice allows Schedule III prescribing (most states permit this for NPs, but some require collaborative agreement specifics)
• Testosterone cypionate/enanthate for injection is the most cost-effective option (generic, widely available)
• Document the diagnostic criteria clearly: two confirmed low morning testosterone levels + symptoms + classification (primary vs. secondary) + absence of contraindications
• Document that reversible causes were addressed (weight loss counseling in obesity, opioid taper consideration, sleep apnea treatment)
• Set expectations with the patient: symptom improvement takes 3–12 months depending on the symptom (libido and energy improve faster than body composition and bone density)
• Schedule follow-up labs at 3 months and 6 months, then annually

Bottom Line

Testosterone testing requires the right collection (morning, fasting, two samples), the right interpretation (consider SHBG in obese/diabetic men), and the right classification (LH/FSH to distinguish primary from secondary). Before prescribing TRT, check PSA, hematocrit, and prolactin; rule out treatable causes like prolactinoma and hemochromatosis; confirm the patient doesn't want fertility; and ensure reversible contributors like obesity and opioids are addressed. Monitor hematocrit religiously—polycythemia is the most common and most dangerous adverse effect. And when a patient arrives already on TRT from an outside clinic, don't panic—just do the workup that should have been done from the start.

Stay sharp out there.