Polycystic ovary syndrome (PCOS) is a common endocrine disorder that affects about 12% of adult women globally.¹ (PCOS was renamed polyendocrine metabolic ovarian syndrome [PMOS]²; however, this Clinical Focus will continue using PCOS until the new name has been widely adopted.) The clinical presentation of PCOS varies substantially, but primary features include androgen excess (eg, hirsutism and acne), reproductive disruption (eg, menstrual irregularity, infertility), and metabolic dysfunction (eg, obesity, insulin resistance, type 2 diabetes).^2,3 The exact causes of PCOS are unknown but are thought to involve complex genetic and environmental interactions. Insulin resistance, accompanied by compensating hyperinsulinemia, is believed to play a key role in PCOS pathophysiology by increasing ovarian and adrenal androgen production.⁴

No single criterion or test can be used to diagnose PCOS. Diagnosis is primarily based on meeting criteria and excluding diseases with similar symptoms. The 2003 Rotterdam criteria are the most-used approach for diagnosis of PCOS and require the presence of at least 2 of 3 criteria in adults: (1) hyperandrogenism, (2) ovulatory dysfunction, and (3) polycystic ovarian morphology (PCOM).³ For adolescents (ages 10-19), hyperandrogenism and ovulatory dysfunction are required for diagnosis.³ Four phenotypes of PCOS are recognized based on different combinations of these criteria (Table 1).⁵

Table 1. Polycystic Ovary Syndrome Phenotypes Based on Rotterdam Criteria [return to contents]

Hyperandrogenism can be diagnosed based on clinical features or biochemical assessment. Features of clinical hyperandrogenism include hirsutism (excess facial or body hair), acne, and female pattern hair loss. To evaluate hirsutism, the modified Ferriman Gallwey score should be used.³ To assess alopecia, visual scales such as Ludwig or Olsen should be considered.³ In addition, diagnosis of clinical hyperandrogenism should take into account natural variations in presentation by ethnicity.³

Ovulatory dysfunction manifests as irregular menstrual cycles and amenorrhea. A diagnosis of PCOS should be considered in individuals with irregular menstrual cycles based on their age and stage of development. The International Evidence-Based Guideline defines irregular menstrual cycles as³

• Normal in the first-year postmenarche as part of the pubertal transition
• >1 to <3 years postmenarche: menstrual cycles <21 days or >45 days
• >3 years postmenarche to perimenopause: menstrual cycles <21 or >35 days or <8 cycles per year
• >1 year postmenarche: >90 days for any 1 cycle
• Primary amenorrhea by age 15 or >3 years after thelarche (beginning of breast development)

Diagnosis of PCOS can be challenging in an adolescent with irregular menstrual cycles. Many menstrual abnormalities of PCOS occur physiologically during puberty (eg, irregular cycles and anovulation). Thus, the value and optimal timing of assessment and diagnosis of PCOS should be discussed with the patient and their parents(s) or guardian(s).³

Either ultrasound or testing for serum anti-Müllerian hormone (AMH) is recommended to identify PCOM in adults.³ To avoid overdiagnosis, only 1 diagnostic method, not both, should be used. PCOM evaluation is not indicated for adolescents since its clinical features overlap with normal adolescent physiology.³

This Clinical Focus discusses the important role that laboratory testing plays in the diagnosis of PCOS based on the Rotterdam criteria and the International Evidence-based Guideline for the assessment and management of PCOS³ as well as its role in the exclusion of other conditions with similar clinical features and assessment of comorbidities. This information is not intended as medical advice. Test selection and interpretation, diagnosis, and patient management decisions should be based on the physician’s education, clinical expertise, and assessment of the patient.

Test availability [return to contents]

Quest Diagnostics offers tests and panels to support diagnosis of PCOS (Table 2) and differential diagnosis of PCOS and conditions with overlapping features (Table 3). Quest also offers tests and panels to diagnose and manage cardiometabolic comorbidities associated with PCOS (Table 4). Panel components may be ordered separately.

Table 2. Laboratory Tests for Assessing PCOS Diagnostic Criteria [return to contents]

Table 3. Laboratory Tests for Differential Diagnosis of PCOS [return to contents]

Table 4. Laboratory Tests for the Diagnosis and Management of Cardiometabolic Comorbidities Associated with PCOS [return to contents]

Test selection and interpretation [return to contents]

The following section summarizes literature and guidelines for test selection and interpretation to support a PCOS diagnosis, distinguish PCOS from conditions with overlapping features, and diagnose and manage comorbidities associated with PCOS.

Diagnosis of PCOS

Laboratory testing can help diagnose PCOS by providing biochemical evidence of hyperandrogenism, assessing ovulatory function, and detecting polycystic ovaries (Figure).^3,9-12

Figure. Suggested Polycystic Ovary Syndrome (PCOS) Diagnostic Algorithm [return to contents]

Hyperandrogenism

Assessment of biochemical hyperandrogenism can help diagnose PCOS when clinical signs of hyperandrogenism (in particular hirsutism) are unclear or absent.³ Testing to assess biochemical hyperandrogenism in the diagnosis of PCOS includes the following:

• Assess for elevated total and free testosterone (test code 36170).³
• If total and free testosterone are normal but PCOS-related hyperandrogenism is highly suspected due to signs such as acne and hirsutism, consider testing for androstenedione (test code 17182) and dehydroepiandrosterone (DHEA) sulfate (test code 402).³
• Concurrent testing of total and free testosterone, DHEA, and androstenedione using the Steroid Panel, Polycystic Ovary Syndrome (PCOS) (test code 90424) can be considered as several studies have demonstrated improved detection of biochemical hyperandrogenism when DHEA/DHEA sulfate and/or androstenedione are included,^13-15 though the value of routinely measuring them is considered low because relatively few additional patients would receive a diagnosis.¹⁶ Concurrent assessment of these secondary androgens with testosterone may provide a more complete biochemical assessment in a single testing visit and reduce the need for repeat testing when initial testosterone results are normal.

The International Evidence-Based Guideline recommends using liquid chromatography tandem mass spectrometry (LC/MS/MS)-based assays for the most accurate assessment of total and free testosterone.³ Measuring total and free testosterone (with LC/MS/MS-based methods) is preferred over measuring total testosterone alone because free testosterone is a more sensitive test for hyperandrogenism.¹⁶ Equilibrium dialysis is the preferred method for determining free testosterone (included in test code 36170),¹⁶ but calculated free testosterone derived from LC/MS/MS-based total testosterone, sex hormone-binding globulin, and albumin (test code 14966) can also be used, though this method is less accurate (Table 2).^3,17 Direct free testosterone assays, such as radiometric or enzyme-linked assays, are not recommended due to limited accuracy in identifying hyperandrogenism in PCOS.³

Ovulatory dysfunction

Ovulatory dysfunction may be present even with regular menstrual cycles. For these patients, anovulation can be confirmed by measuring serum progesterone level (LC/MS, test code 17183; immunoassay, test code 745) in the mid-luteal phase.³

PCOM

Elevated serum AMH (test code 37227) supports a PCOS diagnosis. However, clinicians should be aware of factors that influence AMH levels, including age, body mass index, hormonal contraception, and menstrual cycle day.³ Population- and assay-specific thresholds are recommended for laboratories offering serum AMH testing.³

Differential diagnosis of PCOS

The criteria used to diagnose PCOS overlap with clinical features of other disorders. At minimum, all individuals undergoing evaluation for PCOS should receive testing to exclude nonclassic congenital adrenal hyperplasia (NCCAH), hyperprolactinemia, and thyroid disease.^3,11 Moreover, pregnancy should be ruled out in patients with amenorrhea.¹² Testing for additional causes of clinical features is recommended if indicated by the patient presentation.³ Importantly, PCOS can only be diagnosed once these other disorders have been excluded.³

Laboratory testing can be used for this differential diagnosis (Figure and discussed below). The conditions are listed below in alphabetical order.

Acromegaly

Acromegaly, like PCOS, can have symptoms of oligomenorrhea and hirsutism. However, other suggestive clinical features include changes in facial shape and features (eg, enlarged jaw), headaches, enlarged hands or feet, and peripheral vision impairment.¹¹ Testing for serum insulin-like growth factor-1 (IGF-1, test code 16293) is recommended when acromegaly is suspected, with elevated levels supporting the diagnosis.^11,18

Cushing syndrome

Cushing syndrome has several clinical features that overlap with PCOS (eg, hirsutism, acne, alopecia) as well as more distinctive features such as myopathy and easy bruising.^11,19 Screening for Cushing syndrome is performed by measuring late-night salivary cortisol (LNSC), 24-hour urinary free cortisol, or serum cortisol after dexamethasone-induced suppression.²⁰ Any of these tests, or a combination thereof, are recommended when Cushing syndrome is suspected.²⁰

For LNSC testing, at least 2 measurements are recommended to assess whether the patient exhibits abnormal circadian rhythm of cortisol secretion.²⁰ Quest offers tests that include 1 (test code 19897), 2 (test code 93020), 3 (test code 92687), or 4 (test code 18921) LNSC measurements. For urinary free cortisol testing, at least 2 measurements are recommended to assess for increased bioavailable cortisol (test code 14534 [includes 1 measurement]).²⁰ Lastly, an overnight 1-mg dexamethasone suppression test (test code 6921) is available to evaluate for impaired regulation of cortisol secretion. In the absence of Cushing syndrome, dexamethasone normally inhibits cortisol secretion, and serum cortisol morning levels <1.8 μg/dL indicate normal regulation of cortisol.^20,21 In the presence of Cushing syndrome, elevated serum cortisol morning levels indicate impaired cortisol regulation.^20,21 Testing for appropriately elevated serum dexamethasone (test code 29391) alongside cortisol can assure an adequate dose was administered and help rule out a false-positive result, which can occur due to abnormal absorption of dexamethasone.²⁰ Women taking estrogen-containing oral contraceptives should not receive a dexamethasone suppression test due to elevated risk of a false-positive result.²⁰

Hyperprolactinemia

Hyperprolactinemia can present with ovulatory dysfunction and signs of hyperandrogenism that mimic PCOS.^10,22 It is distinguished from PCOS by high levels of serum prolactin (test code 746). All individuals undergoing evaluation for PCOS should receive serum prolactin testing to rule out hyperprolactinemia.^3,10,11

Hypogonadotropic hypogonadism/functional hypothalamic amenorrhea

Hypogonadotropic hypogonadism, most commonly caused by functional hypothalamic amenorrhea, is suggested by a history of amenorrhea, low body weight, excessive exercise, and a lack of clinical signs of hyperandrogenism.^3,11 If indicated, hypogonadotropic hypogonadism should be excluded by measuring serum luteinizing hormone (LH, test code 615) and serum follicle stimulating hormone (FSH, test code 470).³ FSH and LH may also be ordered together (test code 7137). If functional hypothalamic amenorrhea is specifically suspected, then serum estradiol (test code 30289) should be measured in addition to LH and FSH.^11,12,23

In PCOS, serum FSH levels are typically normal, LH levels are normal or elevated (LH levels are often higher than FSH), and estradiol levels are normal.¹² In hypogonadotropic hypogonadism/functional hypothalamic amenorrhea, FSH and LH are low to low-normal; FSH levels are usually higher than LH. In addition, estradiol levels are typically low in these patients.^11,12,23

NCCAH

NCCAH can be confused with PCOS due to overlapping clinical features such as hirsutism, acne, and oligo/amenorrhea.²⁴ The disease is primarily caused by genetic defects in the enzyme 21-hydroxylase. NCCAH is distinguished from PCOS by increased levels of 17-hydroxyprogesterone (17-OHP) caused by partially diminished 21-hydroxylase activity.²⁴

All individuals undergoing evaluation for PCOS should obtain a serum 17-OHP measurement (test code 17180).^3,10,11 Serum 17-OHP levels (early follicular phase)

• <200 ng/dL exclude NCCAH.⁹
• >1,000 ng/dL indicate NCCAH.⁹
• 200 ng/dL to 1,000 ng/dL indicate consideration of further testing with cosyntropin stimulation (test code 17682; a >1,000 ng/dL response to stimulation suggests NCCAH rather than PCOS).⁹

Pregnancy

For patients presenting with amenorrhea, pregnancy should be excluded during the initial evaluation.^12,23 Pregnancy is distinguished from PCOS by the presence of human chorionic gonadotropin (hCG) detected by testing serum (test code 8396 [quantitative]; 8435 [qualitative]) or urine (test code 396),¹¹ though serum may be preferred due to the risk of false-negative results from urine.¹² However, individuals with circulating heterophilic antibodies (human antibodies directed against animal-derived antigens) can have false-positive serum hCG results due to interference with immunoassay antibodies.^25,26 Specimens can be pretreated to prevent this interference; for this approach, Quest offers hCG testing with pretreatment for human anti-mouse antibodies (HAMA) (test code 19720).

Primary ovarian insufficiency

Primary ovarian insufficiency should be suspected in patients presenting with amenorrhea and symptoms of estrogen deficiency.¹¹ It can be distinguished from PCOS in most patients by low serum levels of estradiol (test code 30289) and elevated serum levels of FSH (test code 470).^11,12

Thyroid disease

Thyroid disease may lead to menstrual disorders similar to those of PCOS and can be distinguished by measuring serum levels of thyroid-stimulating hormone (TSH). The pituitary gland produces more TSH when blood levels of thyroid hormones T3 and T4 are low (hypothyroidism) and less when they are high (hyperthyroidism).²⁷ TSH values (test code 899) above the upper limit of normal typically suggest hypothyroidism, whereas values below the lower limit of normal may suggest hyperthyroidism and the need for further evaluation.¹¹ All individuals undergoing evaluation for PCOS should obtain a measurement of TSH to exclude thyroid disease.^3,10,11

Falsely elevated TSH values may occur in specimens from patients who have circulating heterophilic antibodies, including HAMAs.²⁶ Testing specimens from such patients involves pretreatment to inhibit possible interference (test code 19537).

Virilizing tumors and ovarian hyperthecosis

Virilizing tumors are suggested by rapid progression of hyperandrogenemia symptoms and large elevations in androgens.^3,28 Highly elevated levels of total testosterone (eg, >150-200 ng/dL; test code 15983) increase suspicion of an androgen-producing tumor in the adrenal gland or ovary.^10,28 Additional testing of DHEA sulfate levels (test code 402) can support differential diagnosis, with very high levels (eg, >700 μg/dL) suggesting an adrenal tumor and lower levels suggesting an ovarian tumor.^10,28 However, no specific cutoff levels define the presence of a tumor versus PCOS. Follow-up evaluation using imaging is generally recommended to confirm the presence of a tumor.²⁸

In postmenopausal individuals, severe or worsening hyperandrogenism is suggestive of a virilizing ovarian or adrenal tumor or ovarian hyperthecosis and warrants biochemical and imaging evaluation.³ Elevated levels of total testosterone (test code 15983) with normal levels of adrenal androgens (eg, DHEA sulfate [test code 402]) support a diagnosis of ovarian hyperthecosis, but follow-up imaging is indicated for further evaluation.²⁸

Metabolic dysfunction and comorbidity assessment

The recent name change of PCOS to polyendocrine metabolic ovarian syndrome (PMOS) reflects the growing body of evidence supporting the role that metabolic dysfunction, in particular insulin resistance, plays in both the development and symptomology of the disorder.^4,29-31

PCOS increases risk for metabolic conditions (including insulin resistance, dysglycemia, type 2 diabetes, obesity, dyslipidemia, and metabolic syndrome) as well as cardiovascular disease (CVD).^3,4,10,29,32 All individuals with PCOS should be assessed for metabolic and cardiovascular comorbidities, regardless of weight.³

All patients with PCOS should have their glycemic status assessed, preferably with an oral glucose tolerance test (test code 35181).^3,10 Alternative testing options include fasting plasma glucose (test code 484) and/or glycated hemoglobin (HbA1c) (test code 496). Glycemic status should be reassessed every 1 to 3 years.³

All patients should also be tested with a lipid profile that includes total cholesterol, low-density lipoprotein cholesterol, high-density lipoprotein cholesterol, and triglycerides (test code 7600 or 91716) at diagnosis and at regular intervals thereafter based on overall CVD risk.³ Additional testing may be considered for chronic kidney disease, metabolic dysfunction–associated steatotic liver disease, and other cardiometabolic conditions based on the patient’s clinical history and presentation.

For more information on test selection and interpretation, see related test guides:

• LDL Cholesterol
• Laboratory Testing for Diabetes Diagnosis and Management
• Laboratory Testing for Chronic Kidney Disease Diagnosis and Management