What are the diagnostic steps for primary amenorrhea (absence of menstruation) that lead to a diagnosis of Androgen Insensitivity Syndrome (AIS)?

Diagnostic Steps for Androgen Insensitivity Syndrome in Primary Amenorrhea

The diagnostic pathway for primary amenorrhea leading to Androgen Insensitivity Syndrome (AIS) requires systematic evaluation starting with karyotype testing, hormone analysis, and imaging studies to confirm the presence of testes in a phenotypically female patient.

Initial Evaluation of Primary Amenorrhea

Age-Based Evaluation Timing

• Evaluate at age 13 if no signs of puberty
• Evaluate at age 16 if menarche has not occurred despite normal pubertal development
• Evaluate if menarche has not occurred within 3 years after breast development began 1

History and Physical Examination

• Detailed menstrual history (primary amenorrhea)
• Pubertal development pattern (normal female secondary sexual characteristics)
• Family history of similar presentations
• Physical examination focusing on:
  • Normal female external genitalia
  • Breast development (typically normal or enhanced)
  • Absent or sparse pubic/axillary hair
  • Shortened or blind-ending vagina
  • Absence of cervix on pelvic examination

Laboratory Testing Sequence

1. Pregnancy test (to exclude pregnancy as cause of amenorrhea)

2. Hormone panel:

   • FSH and LH (typically normal or elevated)
   • Testosterone (elevated, in male range)
   • Estradiol (normal female range despite high testosterone)
   • TSH and prolactin (to rule out other causes)
   • LH:FSH ratio (typically <1 in hypothalamic amenorrhea but may be elevated in AIS) 1

3. Karyotype analysis - critical diagnostic test showing 46,XY karyotype in a phenotypically female patient 2, 3

Imaging Studies

1. Pelvic ultrasonography:

   • Absence of uterus and ovaries
   • Possible visualization of testes in inguinal canals 3, 4

2. MRI of pelvis and abdomen if ultrasonography is inconclusive:

   • More accurate localization of gonads (testes)
   • Can detect intra-abdominal testes not visualized on ultrasound
   • Evaluation of Wolffian duct remnants
   • Detection of potential gonadal tumors 5

3. Laparoscopy may be required if imaging is inconclusive for definitive identification of gonads

Genetic Testing

1. Androgen receptor (AR) gene sequencing:

   • Confirms diagnosis by identifying mutations in AR gene on X chromosome
   • Determines whether AIS is complete, partial, or mild form 2

2. SRY gene testing to confirm presence on Y chromosome 6

Confirmatory Testing

1. FISH analysis to confirm presence of AR gene on X chromosome and SRY gene on Y chromosome 6

2. Gonadal biopsy (if needed) to confirm testicular tissue and evaluate for malignancy risk

Differential Diagnosis Considerations

• Müllerian agenesis (46,XX karyotype with normal ovarian function)
• 5-alpha reductase deficiency (may have more ambiguous genitalia)
• 17-beta hydroxysteroid dehydrogenase deficiency
• Congenital adrenal hyperplasia (typically with signs of virilization) 7
• Hypothalamic amenorrhea (low LH, FSH, estradiol with female karyotype) 1

Management After Diagnosis

• Multidisciplinary team approach including endocrinology, gynecology, genetics, and psychology
• Gonadectomy recommended after puberty due to malignancy risk (5-10%)
• Estrogen replacement therapy
• Vaginal dilation or surgical procedures for vaginal hypoplasia
• Psychological support for gender identity and fertility counseling 2, 4

Common Pitfalls to Avoid

• Misdiagnosis as PCOS or hypothalamic amenorrhea
• Delayed karyotype testing in patients with primary amenorrhea
• Failure to locate gonads with ultrasound alone (MRI superior for intra-abdominal testes) 5
• Delayed gonadectomy increasing risk of malignancy 4, 6
• Inadequate psychological support during and after diagnosis

The diagnostic approach to AIS requires systematic evaluation with karyotype testing being the pivotal diagnostic step that should be performed in all cases of primary amenorrhea with normal female secondary sexual characteristics but absence of uterus on imaging.