What tests should be run for a 24-year-old female with amenorrhea (absence of menstruation) and breast tenderness, with a previously negative Human Chorionic Gonadotropin (HCG) test?

Diagnostic Workup for Amenorrhea with Breast Tenderness and Prior Negative Pregnancy Test

Repeat Pregnancy Testing First

The first and most critical test is to repeat a quantitative serum hCG (beta-hCG) immediately, as pregnancy remains the most common cause of amenorrhea and a negative test "last month" does not exclude current pregnancy. 1, 2, 3, 4

• A urine pregnancy test from last month may have been performed too early to detect pregnancy, as most qualitative tests require an additional 11 days past expected menses to detect 100% of pregnancies 1
• Serum quantitative hCG is more sensitive and reliable than urine testing, particularly in early pregnancy when levels may be below urine test detection thresholds 1
• If the serum hCG is positive but low, obtain repeat measurement in 48 hours to assess for appropriate doubling, which helps distinguish viable intrauterine pregnancy from ectopic pregnancy or early pregnancy loss 1

Initial Hormone Panel

After confirming pregnancy status, obtain the following serum tests simultaneously 2, 3, 4:

• Thyroid-stimulating hormone (TSH) - to evaluate for thyroid dysfunction, which commonly causes amenorrhea 2, 3, 4
• Prolactin - elevated levels suggest hyperprolactinemia, which can cause both amenorrhea and breast tenderness 2, 3, 4
• Follicle-stimulating hormone (FSH) - helps distinguish ovarian failure from hypothalamic/pituitary dysfunction 2, 3, 4
• Luteinizing hormone (LH) - the FSH:LH ratio helps identify polycystic ovary syndrome (PCOS) 2, 4

Clinical Context for Breast Tenderness

The breast tenderness in this case warrants specific consideration 5:

• Cyclical mastalgia (related to hormonal fluctuations) is typically bilateral and suggests functioning ovarian tissue, making primary ovarian insufficiency less likely 5
• Two weeks of breast tenderness could represent early pregnancy symptoms, further emphasizing the need for repeat pregnancy testing 5
• If prolactin is elevated, this can directly cause breast tenderness and galactorrhea 2, 3

Algorithmic Approach Based on Initial Results

If hCG is Positive:

• Obtain transvaginal ultrasound when hCG reaches 1,000-3,000 mIU/mL to confirm intrauterine pregnancy location 1
• Serial hCG measurements every 48 hours if initial level is low to assess viability 1

If Prolactin is Elevated (>25 ng/mL):

• Order brain MRI to evaluate for pituitary adenoma 2, 3
• Assess for medications causing hyperprolactinemia (antipsychotics, metoclopramide) 2

If TSH is Abnormal:

• Add free T4 to characterize thyroid dysfunction 2, 4
• Both hypothyroidism and hyperthyroidism can cause amenorrhea 2

If FSH is Elevated (>40 mIU/mL):

• Suggests primary ovarian insufficiency 2, 4
• Repeat FSH in 1 month with estradiol level for confirmation 4
• Consider karyotype if patient is under 30 years old to evaluate for Turner syndrome or variants 2, 4

If All Initial Tests are Normal:

• Perform progesterone challenge test (medroxyprogesterone acetate 10 mg daily for 10 days) to assess estrogen status and outflow tract patency 3
• Withdrawal bleeding indicates adequate estrogen and patent outflow tract, suggesting anovulation (likely PCOS) 3
• No withdrawal bleeding suggests either outflow tract obstruction or hypoestrogenic state requiring further evaluation 3

Critical Pitfalls to Avoid

• Never assume a negative pregnancy test from weeks ago excludes current pregnancy - hCG levels rise rapidly in early pregnancy, and testing timing is critical 1, 2
• Do not dismiss breast tenderness as benign without proper evaluation - while rarely indicating malignancy, it provides important diagnostic clues about hormonal status 5
• Avoid ordering extensive imaging before obtaining basic hormone levels - the history, exam, and initial labs will guide whether pelvic ultrasound or brain MRI is needed 2, 4
• Remember that patients with primary ovarian insufficiency can maintain unpredictable ovarian function - they should not be presumed infertile and may still require contraception 2, 4

Additional Considerations Based on History

If the patient has 4:

• Weight loss, excessive exercise, or stress - consider functional hypothalamic amenorrhea and evaluate for eating disorders and low bone density 2, 4
• Hirsutism, acne, or obesity - increases likelihood of PCOS; add testosterone and DHEA-S to initial panel 2, 4
• Galactorrhea - strongly supports prolactin testing and subsequent pituitary imaging if elevated 2, 3
• Vasomotor symptoms (hot flashes) - suggests hypoestrogenic state from ovarian insufficiency 4