Test Catalog

Test ID: CYPZ    
21-Hydroxylase Gene (CYP21A2), Full Gene Analysis, Varies

Useful For Suggests clinical disorders or settings where the test may be helpful

Carrier screening and diagnosis of 21-hydroxylase deficient congenital adrenal hyperplasia (CAH) in individuals with a personal or family history of 21-hydroxylase deficiency, or as follow-up to positive CAH newborn screens and/or measurement of basal and adrenocorticotropic hormone- 1-24 stimulated 17-hydroxyprogesterone, androstenedione, and other adrenal steroid levels


May be used to identify CYP21A2 mutations in individuals with a suspected diagnosis of 21-hydroxylase deficient CAH when a common mutation panel is negative or only identifies 1 mutation.


In prenatal cases of ambiguous genitalia detected by ultrasound, particularly when the fetus is confirmed XX female by chromosome analysis.


This test ID should also be used for known/familial variant analysis for CYP21A2. Due to the complexity of the CYP21A2 locus, site specific testing for known/familial variants is not offered for this gene.

Genetics Test Information Provides information that may help with selection of the correct genetic test or proper submission of the test request

This test includes Sanger gene sequencing and multiplex ligation-dependent probe amplification to evaluate the CYP21A2 gene for carrier screening and diagnosis of 21-hydroxylase deficient congenital adrenal hyperplasia (CAH).

Testing Algorithm Delineates situations when tests are added to the initial order. This includes reflex and additional tests.

For prenatal specimens only: If amniotic fluid (nonconfluent cultured cells) is received, amniotic fluid culture will be added and charged separately. If chorionic villus specimen (nonconfluent cultured cells) is received, fibroblast culture will be added and charged separately. For any prenatal specimen that is received, maternal cell contamination studies will be added.

Clinical Information Discusses physiology, pathophysiology, and general clinical aspects, as they relate to a laboratory test

Congenital adrenal hyperplasia (CAH), with an incidence rate of 1 in 10,000 to 18,000 live births, is one of the most common inherited syndromes. The condition is characterized by impaired cortisol production due to inherited defects in steroid biosynthesis. The clinical consequences of CAH, besides diminished cortisol production, depend on which enzyme is affected and whether the loss of function is partial or complete.


In greater than 90% of CAH cases, the affected enzyme is 21-steroid hydroxylase, encoded by the CYP21A2 gene located on chromosome 6 within the highly recombinant human histocompatibility complex locus. 21-hydroxylase deficient CAH is inherited in an autosomal recessive pattern and has a spectrum of clinical phenotypes depending upon residual enzyme activity. Excessive adrenal androgen biosynthesis results in varying degrees of virilization. If there is some residual enzyme activity, a non-classical phenotype results, with signs of hyperandrogenism typically starting in later childhood or adolescence. Individuals with severe enzyme deficiency have classical CAH, with prenatal onset of virilization. Classical CAH which is subdivided into simple-virilizing (minimal residual enzyme activity) and salt-wasting (no residual enzyme activity) forms. Patients with salt-wasting CAH have both cortisol and mineral corticosteroid deficiency and are at risk for life-threatening salt-wasting crises if untreated.


Because of its high incidence rate, 21-hydroxylase deficiency is screened for in most US newborn screening programs, typically by measuring 17-hydroxyprogesterone concentrations in blood spots by immunoassay. Confirmation by other testing strategies (eg, LC-MS/MS, CAHBS / Congenital Adrenal Hyperplasia [CAH] Newborn Screening, Blood Spot), or retesting after several weeks, is required for most positive screens because of the high false-positive rates of the immunoassays (due to physiological elevations of 17-hydroxyprogesterone in premature babies and immunoassay cross-reactivity with other steroids). In a small percentage of cases, additional testing will fail to provide a definitive diagnosis. In addition, screening strategies can miss many non-classical cases, which may present later in childhood or adolescence and require more extensive steroid hormone profiling, including testing before and after adrenal stimulation with adrenocorticotropic hormone (ACTH)-1-24.


For these reasons, genetic diagnosis plays an important ancillary role in both classical and non-classical cases. In addition, the high carrier frequency (approximately 1 in 50) for CYP21A2 mutations makes genetic diagnosis important for genetic counseling. Genetic testing can also play a role in prenatal diagnosis of 21-hydroxylase deficiency. However, accurate genetic diagnosis continues to be a challenge because most of the mutations arise from recombination events between CYP21A2 and its highly homologous pseudogene, CYP21A1P (transcriptionally inactive). In particular, partial or complex rearrangements (with or without accompanying gene duplication events), which lead to reciprocal exchanges between gene and pseudogene, can present severe diagnostic challenges. Comprehensive genetic testing strategies must therefore allow accurate assessment of most, or all, known rearrangements and mutations, as well as unequivocal determination of whether the observed changes are located within a potentially transcriptionally active genetic segment. Testing of additional family members is often needed for clarification of genetic test results.

Reference Values Describes reference intervals and additional information for interpretation of test results. May include intervals based on age and sex when appropriate. Intervals are Mayo-derived, unless otherwise designated. If an interpretive report is provided, the reference value field will state this.

An interpretive report will be provided.

Interpretation Provides information to assist in interpretation of the test results

All detected alterations will be evaluated according to American College of Medical Genetics and Genomics (ACMG) recommendations. Variants will be classified based on known, predicted, or possible pathogenicity and reported with interpretive comments detailing their potential or known significance.

Cautions Discusses conditions that may cause diagnostic confusion, including improper specimen collection and handling, inappropriate test selection, and interfering substances

Because of the complexity of the genetic structure of the CYP21A2 locus, and the possibility that a patient's congenital adrenal hyperplasia (CAH) may be due to other gene defects, genetic testing results should be correlated carefully with clinical and biochemical data.


This testing strategy is superior to approaches previously used, but may still miss some complex and large-scale genetic rearrangements or deletions, as well as genetic changes in far upstream or downstream gene-regulatory elements that impair CYP21A2 gene expression. This can lead to false-negative test results.


Rare polymorphisms in primer binding sites can lead to selective allelic drop-out, which can lead to false-negative or false-positive diagnosis.


Patients without genetic evidence for disease-causing CYP21A2 genetic changes may still have CAH, but due to a different enzyme defect. Additional and expanded biochemical steroid profiling is, therefore, recommended if the clinical picture is strongly suggestive of CAH.

Clinical Reference Recommendations for in-depth reading of a clinical nature

1. Richards S, Aziz N, Bale S, et al: Standards and guidelines for the interpretation of sequence variants: a joint consensus recommendation of the American College of Medical Genetics and Genomics and the Association for Molecular Pathology. Genet Med 2015 May;17(5):405-424

2. Collett-Solberg PF: Congenital adrenal hyperplasias: from clinical genetics and biochemistry to clinical practice, part I. Clin Pediatr 2001;40:1-16

3. Mercke DP, Bornstein SR, Avila NA, Chrousos GP: NIH conference: future directions in the study and management of congenital adrenal hyperplasia due to 21-hydroxylase deficiency. Ann Intern Med 2002;136:320-334

4. Speiser PW, White PC: Medical progress: congenital adrenal hyperplasia. N Engl J Med 2003;349:776-788

Special Instructions Library of PDFs including pertinent information and forms related to the test