Unfortunately, the number of differentials to consider for PAIS is particularly large. Prompt diagnosis is particularly urgent when a child is born with ambiguous genitalia, as some causes are associated with potentially life-threatening adrenal crises. Determination of testosterone, testosterone precursors and dihydrotestosterone (DHT) at baseline and / or after human chorionic gonadotropin (hCG) stimulation can be used to exclude such defects in androgen biosynthesis.

Approximately one half of all 46,XY individuals born with ambiguous genitalia will not receive a definitive diagnosis. Androgen receptor (AR) gene mutations cannot be found in 27% to 72% of individuals with PAIS. As a result, genetic analysis can be used to confirm a diagnosis of PAIS, but it cannot be used to rule out PAIS. Evidence of abnormal androgen binding in a genital skin fibroblast study has long been the gold standard for the diagnosis of PAIS, even when an AR mutation is not present. However, some cases of PAIS, including AR-mutant-positive cases, will show normal androgen binding. A family history consistent with X-linked inheritance is more commonly found in AR-mutant-positive cases than AR-mutant-negative cases.

The use of dynamic endocrine tests is particularly helpful in isolating a diagnosis of PAIS. One such test is the human chorionic gonadotropin (hCG) stimulation test. If the gonads are testes, there will be an increase in the level of serum testosterone in response to the hCG, regardless of testicular descent. The magnitude of the testosterone increase can help differentiate between androgen resistance and gonadal dysgenesis, as does evidence of a uterus on ultrasound examination. Testicular function can also be assessed by measuring serum anti-Müllerian hormone levels, which in turn can further differentiate PAIS from gonadal dysgenesis and bilateral anorchia.

Another useful dynamic test involves measuring the response to exogenous steroids; individuals with AIS show a decreased response in serum sex hormone binding globulin (SHBG) after a short term administration of anabolic steroids. Two studies indicate that measuring the response in SHBG after the administration of stanozolol could help to differentiate individuals with PAIS from those with other causes of ambiguous genitalia, although the response in individuals with predominantly male phenotypes overlaps somewhat with the response in normal males.

CAIS can only be diagnosed in normal phenotypic females. It is not usually suspected unless the menses fail to develop at puberty, or an inguinal hernia presents during premenarche. As many as 1–2% of prepubertal girls that present with an inguinal hernia will also have CAIS.

A diagnosis of CAIS or Swyer syndrome can be made in utero by comparing a karyotype obtained by amniocentesis with the external genitalia of the fetus during a prenatal ultrasound. Many infants with CAIS do not experience the normal, spontaneous neonatal testosterone surge, a fact which can be diagnostically exploited by obtaining baseline luteinizing hormone and testosterone measurements, followed by a human chorionic gonadotropin (hGC) stimulation test.

The main differentials for CAIS are complete gonadal dysgenesis (Swyer syndrome) and Müllerian agenesis (Mayer-Rokitansky-Kuster-Hauser syndrome or MRKH). Both CAIS and Swyer syndrome are associated with a 46,XY karyotype, whereas MRKH is not; MRKH can thus be ruled out by checking for the presence of a Y chromosome, which can be done either by fluorescence in situ hybridization (FISH) analysis or on full karyotype. Swyer syndrome is distinguished by poor breast development and shorter stature. The diagnosis of CAIS is confirmed when androgen receptor (AR) gene sequencing reveals a mutation, although up to 5% of individuals with CAIS do not have an AR mutation.

Up until the 1990s, a CAIS diagnosis was often hidden from the affected individual and / or family. It is current practice to disclose the genotype at the time of diagnosis, particularly when the affected girl is at least of adolescent age. If the affected individual is a child or infant, it is generally up to the parents, often in conjunction with a psychologist, to decide when to disclose the diagnosis.

Individuals with CAIS are raised as females. They are born phenotypically female and almost always have a heterosexual female gender identity; the incidence of homosexuality in women with CAIS is thought to be less than unaffected women. However, at least two case studies have reported male gender identity in individuals with CAIS.

MAIS is only diagnosed in normal phenotypic males, and is not typically investigated except in cases of male infertility. MAIS has a mild presentation that often goes unnoticed and untreated; even with semenological, clinical and laboratory data, it can be difficult to distinguish between men with and without MAIS, and thus a diagnosis of MAIS is not usually made without confirmation of an AR gene mutation. The androgen sensitivity index (ASI), defined as the product of luteinizing hormone (LH) and testosterone (T), is frequently raised in individuals with all forms of AIS, including MAIS, although many individuals with MAIS have an ASI in the normal range. Testosterone levels may be elevated despite normal levels of luteinizing hormone. Conversion of testosterone (T) to dihydrotestosterone (DHT) may be impaired, although to a lesser extent than is seen in 5α-reductase deficiency. A high ASI in a normal phenotypic male, especially when combined with azoospermia or oligospermia, decreased secondary terminal hair, and/or impaired conversion of T to DHT, can be indicative of MAIS, and may warrant genetic testing.

Gonadectomy at time of diagnosis is the current recommendation for PAIS if presenting with cryptorchidism, due to the high (50%) risk of germ cell malignancy. The risk of malignancy when testes are located intrascrotally is unknown; the current recommendation is to biopsy the testes at puberty, allowing investigation of at least 30 seminiferous tubules, with diagnosis preferably based on OCT3/4 immunohistochemistry, followed by regular examinations. Hormone replacement therapy is required after gonadectomy, and should be modulated over time to replicate the hormone levels naturally present in the body during the various stages of puberty. Artificially induced puberty results in the same, normal development of secondary sexual characteristics, growth spurt, and bone mineral accumulation. Women with PAIS may have a tendency towards bone mineralization deficiency, although this increase is thought to be less than is typically seen in CAIS, and is similarly managed.

In cases where the individual is being evaluated for ambiguous genitalia, such as a small phallus, hypospadias, or labioscrotal folds, exploratory surgery may be used to determine if male and/or female internal genitalia is present.

A standard karyotype can be completed to cytogenetically determine that an individual with a partial or complete male phenotype has a XX genotype.

FISH analysis determines the presence or absence of the SRY gene.

Localization of the SRY gene can by determined using fluorescent "in situ" hybridization.

Indicators include two testes which have not descended the inguinal canal, although this is seen in a minority of XX males, and the absence of Müllerian tissue.

Low testosterone can be identified through a simple blood test performed by a laboratory, ordered by a health care provider. Blood for the test must be taken in the morning hours, when levels are highest, as levels can drop by as much as 13% during the day and all normal reference ranges are based on morning levels. However, low testosterone in the absence of any symptoms does not clearly need to be treated.

Normal total testosterone levels depend on the man's age but generally range from 240–950 ng/dL (nanograms per deciliter) or 8.3-32.9 nmol/L (nanomoles per liter). Some men with normal total testosterone have low free or bioavailable testosterone levels which could still account for their symptoms. Men with low serum testosterone levels should have other hormones checked, particularly luteinizing hormone to help determine why their testosterone levels are low and help choose the most appropriate treatment (most notably, testosterone is usually not appropriate for secondary or tertiary forms of male hypogonadism, in which the LH levels are usually reduced).

Treatment is often prescribed for total testosterone levels below 230 ng/dL with symptoms. If the serum total testosterone level is between 230 and 350 ng/dL, free or bioavailable testosterone should be checked as they are frequently low when the total is marginal.

The standard range given is based off widely varying ages and, given that testosterone levels naturally decrease as humans age, age-group specific averages should be taken into consideration when discussing treatment between doctor and patient. In men, testosterone falls approximately 1 to 3 percent each year.

- Blood testing

A position statement by the Endocrine Society expressed dissatisfaction with most assays for total, free, and bioavailable testosterone. In particular, research has questioned the validity of commonly administered assays of free testosterone by radioimmunoassay. The free androgen index, essentially a calculation based on total testosterone and sex hormone-binding globulin levels, has been found to be the worst predictor of free testosterone levels and should not be used. Measurement by equilibrium dialysis or mass spectroscopy is generally required for accurately results, particularly for free testosterone which is present normal in such small concentrations.

Management of AIS is currently limited to symptomatic management; no method is currently available to correct the malfunctioning androgen receptor proteins produced by "AR" gene mutations. Areas of management include sex assignment, genitoplasty, gonadectomy in relation to tumor risk, hormone replacement therapy, genetic counseling, and psychological counseling.

The diagnosis is often one of exclusion found during the workup of delayed puberty.

A paper published in 2012 by Prof. Jacques Young highlights a typical example of the diagnostic work up involved in a suspected case of KS/CHH.

One of the biggest problems in the diagnosis of KS and other forms of CHH is the ability to distinguish between a normal constitutional delay of puberty and KS or CHH.

The main biochemical parameters in men are low serum testosterone and low levels of the gonadotropins LH and FSH, and in women low serum oestrogen and low levels of LH and FSH.

For both males and females with constitutional delay of puberty, endogenous puberty will eventually commence without treatment. However a delay in treatment in a case of KS/HH will delay the physical development of the patient and can cause severe psychological damage. The "wait and see" approach applied to "late bloomers" is probably counterproductive to the needs of the patient whereas a step-by-step approach with hormone replacement therapy used with slowly increasing doses can be used as a diagnostic tool.

Post natal diagnosis of KS / CHH before the age of 6 months is sometimes possible. The normal post natal hormonal surge of gonadotropins along with testosterone or oestrogen is absent in babies with KS / CHH. This lack of detectable hormones in the blood can be used as a diagnostic indicator, especially in male infants.

Normally testicular enlargement is the key sign for the onset of puberty in boys however the use of nighttime LH sampling can help predict the onset of puberty.

In females diagnosis is sometimes further delayed as other causes of amenorrhoea normally have to be investigated first before a case of KS/CHH is considered. KS/CHH can still occur in females in cases when menstruation has begun but stopped after one or two menstrual bleeds. A study of GnRH deficient women in 2011 showed that 10% had experienced one or two bleeds before the onset of amenorrhoea.

In males, treatment with age-appropriate levels of testosterone can be used to distinguish between a case of KS/CHH from a case of delayed puberty. If just delayed the testosterone can "kick-start" endogenous puberty, as demonstrated by testicular enlargement, whereas in the case of KS/CHH there will be no testicular enlargement while on testosterone therapy alone. If no puberty is apparent, especially no testicular development, then a review by a reproductive endocrinologist may be appropriate. Dr Richard Quinton, a leading UK expert on KS/CHH, suggests that if puberty is not apparent by the age of 16 then the patient should be referred for endocrinological review.

A full endocrine workup will be required to measure the levels of the other pituitary hormones, especially prolactin, to check that the pituitary gland is working correctly. There can be other general health issues such as being overweight or having an underlying chronic or acute illness which could cause a delay of puberty. This makes it essential for a patient to get a full endocrine review to distinguish between a case of KS/CHH and another cause for the pubertal delay.

Bone age can be assessed using hand and wrist X-rays. If the bone age is significantly lower than the chronological age of the patient, this could suggest delayed puberty unless there is another underlying reason for the discrepancy.

A karyotype may be performed to rule out Klinefelter syndrome and Turner syndrome, although the hormones levels would also rule out both these relatively common reasons for hypogonadism.

A magnetic resonance imaging (MRI) scan can be used to determine whether the olfactory bulb is present and to check for any physical irregularities of the pituitary gland or hypothalamus.

A standard smell test can be used to check for anosmia, but it must be remembered that even in total anosmia various substances (such as menthol and alcohol) can still be detected by direct stimulation of the trigeminal nerve.

Genetic screening can be carried out, but in light of the unknown genes involved in the majority of KS and CHH cases, a negative result will not rule out a possible diagnosis.

A review paper published in 2014 highlighted the need for doctors to be aware of the possible diagnosis of KS / HH if pubertal delay is found alongside associated "red flag" symptoms. The symptoms listed in the paper were split into two categories; reproductive symptoms associated with the lack of mini puberty seen between birth and six months of age and non-reproductive symptoms which are associated with specific forms of HH. As with other review papers the authors also warned against the "wait and see" approach when puberty appears to be delayed.

Since the Sertoli cells are not affected by Leydig cell hypoplasia, anti-Müllerian hormone is secreted normally and so there are no Müllerian structures. Wolffian structures, such as the prostate, vasa deferentia, and epidydimides are present. In type I, abdominal testes are revealed on ultrasound; in type II testes may be descended or undescended.

People with Leydig cell hypoplasia type I display no response to the hCG stimulation test; there is no increase in serum levels of testosterone and dihydrotestosterone. Leydig cell hypoplasia type II can display either a pronounced rise of testosterone levels or no rise.

In any case, the diagnosis is confirmed on biopsy of the testes, revealing either absent or hypoplastic Leydig cells. The inside of the testis will be grayish and mucous, displaying arrested spermatogenesis and the presence of Sertoli cells. The diagnosis can also be confirmed by looking for mutations in the gene for the LH receptor.

A diagnosis of Leydig cell hypoplasia is usually made in the neonatal period, following the discovery of ambiguous genitalia, or at puberty, when secondary sex characteristics fail to develop. Puberty is the most common time for Leydig cell hypoplasia to be diagnosed.

Patients with Leydig cell hypoplasia may be treated with hormone replacement therapy (i.e., with androgens), which will result in normal sexual development and the resolution of most symptoms. In the case of 46,XY (genetically "male") individuals who are phenotypically female and/or identify as the female gender, estrogens should be given instead. Surgical correction of the genitals in 46,XY males may be required, and, if necessary, an orchidopexy (relocation of the undescended testes to the scrotum) may be performed as well.

Conditions justifying newborn screening for any disorder include (1) a simple test with an acceptable sensitivity and specificity, (2) a dire consequence if not diagnosed early, (3) an effective treatment if diagnosed, and (4) a frequency in the population high enough to justify the expense. In the last decade more states and countries are adopting newborn screening for salt-wasting CAH due to 21-hydroxylase deficiency, which leads to death in the first month of life if not recognized.

The salt-wasting form of CAH has an incidence of 1 in 15,000 births and is potentially fatal within a month if untreated. Steroid replacement is a simple, effective treatment. However, the screening test itself is less than perfect. While the 17α-hydroxyprogesterone level is easy to measure and sensitive (rarely missing real cases), the test has a poorer specificity. Screening programs in the United States have reported that 99% of positive screens turn out to be false positives upon investigation of the infant. This is a higher rate of false positives than the screening tests for many other congenital metabolic diseases.

When a positive result is detected, the infant must be referred to a pediatric endocrinologist to confirm or disprove the diagnosis. Since most infants with salt-wasting CAH become critically ill by 2 weeks of age, the evaluation must be done rapidly despite the high false positive rate.

Levels of 17α-hydroxyprogesterone, androstenedione, and cortisol may play a role in screening.

Testing serum LH and FSH levels are often used to assess hypogonadism in women, particularly when menopause is believed to be happening. These levels change during a woman's normal menstrual cycle, so the history of having ceased menstruation coupled with high levels aids the diagnosis of being menopausal. Commonly, the post-menopausal woman is not called hypogonadal if she is of typical menopausal age. Contrast with a young woman or teen, who would have hypogonadism rather than menopause. This is because hypogonadism is an abnormality, whereas menopause is a normal change in hormone levels. In any case, the LH and FSH levels will rise in cases of primary hypogonadism or menopause, while they will be low in women with secondary or tertiary hypogonadism.

Hypogonadism is often discovered during evaluation of delayed puberty, but ordinary delay, which eventually results in normal pubertal development, wherein reproductive function is termed constitutional delay. It may be discovered during an infertility evaluation in either men or women.

Preimplantation genetic diagnosis (PGD or PIGD) refers to genetic profiling of embryos prior to implantation (as a form of embryo profiling), and sometimes even of oocytes prior to fertilization. When used to screen for a specific genetic sequence, its main advantage is that it avoids selective pregnancy termination, as the method makes it highly likely that a selected embryo will be free of the condition under consideration.

In the UK, AIS appears on a list of serious genetic diseases that may be screened for via PGD. Some ethicists, clinicians, and intersex advocates have argued that screening embryos to specifically exclude intersex traits are based on social and cultural norms as opposed to medical necessity.

Due to the inability of the streak gonads to produce sex hormones (both estrogens and androgens), most of the secondary sex characteristics do not develop. This is especially true of estrogenic changes such as breast development, widening of the pelvis and hips, and menstrual periods. As the adrenal glands can make limited amounts of androgens and are not affected by this syndrome, most of these persons will develop pubic hair, though it often remains sparse.

Evaluation of delayed puberty usually reveals elevation of gonadotropins, indicating that the pituitary is providing the signal for puberty but the gonads are failing to respond. The next steps of the evaluation usually include checking a karyotype and imaging of the pelvis. The karyotype reveals XY chromosomes and the imaging demonstrates the presence of a uterus but no ovaries (the streak gonads are not usually seen by most imaging). Although an XY karyotype can also indicate a person with complete androgen insensitivity syndrome, the absence of breasts, and the presence of a uterus and pubic hair exclude the possibility. At this point it is usually possible for a physician to make a diagnosis of Swyer syndrome.

The consequences to the girl with XX gonadal dysgenesis:

1. Her gonads cannot make estrogen, so her breasts will not develop and her uterus will not grow and menstruate until she is given estrogen. This is often given through the skin now.

2. Her gonads cannot make progesterone, so her menstrual periods will not be predictable until she is given a progestin, still usually as a pill.

3. Her gonads cannot produce eggs so she will not be able to conceive children naturally. A woman with a uterus but no ovaries may be able to become pregnant by implantation of another woman's fertilized egg (embryo transfer).

Due to its mild presentation, MAIS often goes unnoticed and untreated. Management of MAIS is currently limited to symptomatic management; methods to correct a malfunctioning androgen receptor protein that result from an AR gene mutation are not currently available. Treatment includes surgical correction of mild gynecomastia, minor hypospadias repair, and testosterone supplementation. Supraphysiological doses of testosterone have been shown to correct diminished secondary sexual characteristics in men with MAIS, as well as to reverse infertility due to low sperm count. As is the case with PAIS, men with MAIS will experience side effects from androgen therapy (such as the suppression of the hypothalamic-pituitary-gonadal axis) at a higher dosage than unaffected men. Careful monitoring is required to ensure the safety and efficacy of treatment. Regular breast and prostate examinations may be necessary due to comorbid association with breast and prostate cancers.

Since CAH is an autosomal recessive disease, most children with CAH are born to parents unaware of the risk and with no family history. Each child will have a 25% chance of being born with the disease. Families typically wish to minimize the degree of virilization of a girl. There is no known prenatal harm to a male fetus from CAH, so treatment can begin at birth.

Adrenal glands of female fetuses with CAH begin producing excess testosterone by the 9th week of gestation. The most important aspects of virilization (urogenital closure and phallic urethra) occur between 8 and 12 weeks. Theoretically, if enough glucocorticoid could be supplied to the fetus to reduce adrenal testosterone production by the 9th week, virilization could be prevented and the difficult decision about timing of surgery avoided.

The challenge of preventing severe virilization of girls is twofold: detection of CAH at the beginning of the pregnancy, and delivery of an effective amount of glucocorticoid to the fetus without causing harm to the mother.

The first problem has not yet been entirely solved, but it has been shown that if dexamethasone is taken by a pregnant woman, enough can cross the placenta to suppress fetal adrenal function.

At present no program screens for risk in families who have not yet had a child with CAH. For families desiring to avoid virilization of a second child, the current strategy is to start dexamethasone as soon as a pregnancy has been confirmed even though at that point the chance that the pregnancy is a girl with CAH is only 12.5%. Dexamethasone is taken by the mother each day until it can be safely determined whether she is carrying an affected girl.

Whether the fetus is an affected girl can be determined by chorionic villus sampling at 9–11 weeks of gestation, or by amniocentesis at 15–18 weeks gestation. In each case the fetal sex can be determined quickly, and if the fetus is a male the dexamethasone can be discontinued. If female, fetal DNA is analyzed to see if she carries one of the known abnormal alleles of the "CYP21" gene. If so, dexamethasone is continued for the remainder of the pregnancy at a dose of about 1 mg daily.

Most mothers who have followed this treatment plan have experienced at least mild cushingoid effects from the glucocorticoid but have borne daughters whose genitalia are much less virilized.

Swyer syndrome represents one phenotypic result of a failure of the gonads to develop properly, and hence is part of a class of conditions termed gonadal dysgenesis. There are many forms of gonadal dysgenesis.

Swyer syndrome is an example of a condition in which an externally unambiguous female body carries dysgenetic, atypical, or abnormal gonads. Other examples include complete androgen insensitivity syndrome, partial X chromosome deletions, lipoid congenital adrenal hyperplasia, and Turner syndrome.

Identification of 45,X/46,XY karyotype has significant clinical implications due to known effects on growth, hormonal balance, gonadal development and histology. 45,X/46,XY is diagnosed by examining the chromosomes in a blood sample.

The age of diagnosis varies depending on manifestations of disease prompting reason for cytogenetic testing. Many patients are diagnosed prenatally due to fetal factors (increased nuchal fold, or abnormal levels of serum), maternal age or abnormal ultrasounds, while others will be diagnosed postnatal due to external genital malformation. It is not uncommon for patients to be diagnosed later in life due to short stature or delayed puberty, or a combination of both.

45,X/46,XY mosaicism can be detected prenatally through amniocentesis however, it was determined that the proportion of 45,X cells in the amniotic fluid cannot predict any phenotypic outcomes, often making prenatal genetic counselling difficult.

Reversal of symptoms have been reported in between 15% to 22% of cases. The causes of this reversal are still under investigation but have been reported in both males and females.

Reversal appears to be associated with 14 of the known gene defects linked to KS/CHH. The study suggests no obvious gene defect showing a tendency to allow reversal. There is a suggestion that the TAC3 and TACR3 mutations might allow for a slightly higher chance of reversal, but the numbers involved are too low to confirm this. The ANOS1 mutations appear to be least likely to allow reversal with to date only one recorded instance in medical literature. Even male patients who previous had micro-phallus or cryptorchidism have been shown to undergo reversal of symptoms.

The reversal might not be permanent and remission can occur at any stage; the paper suggests that this could be linked to stress levels. The paper highlighted a reversal case that went into remission but subsequently achieved reversal again, strongly suggesting an environmental link.

Reversal cases have been seen in cases of both KS and normosmic CHH but appear to be less common in cases of KS (where the sense of smell is also affected). A paper published in 2016 agreed with the theory that there is a strong environmental or epigenetic link to the reversal cases. The precise mechanism of reversal is unclear and is an area of active research.

Reversal would be apparent if testicular development was seen in men while on testosterone therapy alone or in women who menstruate or achieved pregnancy while on no treatment. To date there have been no recorded cases of the reversal of anosmia found in Kallmann syndrome cases.

In terms of the diagnosis of 17β-hydroxysteroid dehydrogenase III deficiency the following should be taken into account:

- Delta(4)-A to T ratio (unusually increased)

- Thyroid dyshormonogenesis

- Genetic testing

The 2006 Consensus statement on the management of intersex disorders states that individuals with 17β-hydroxysteroid dehydrogenase III deficiency have an intermediate risk of germ cell malignancy, at 28%, recommending that gonads be monitored. A 2010 review put the risk of germ cell tumors at 17%.

The management of 17β-hydroxysteroid dehydrogenase III deficiency can consist, according to one source, of the elimination of gonads prior to puberty, in turn halting masculinization.

Hewitt and Warne state that, children with 17β-hydroxysteroid dehydrogenase III deficiency who are raised as girls often later identify as male, describing a "well known, spontaneous change of gender identity from female to male" that "occurs after the onset of puberty." A 2005 systematic review of gender role change identified the rate of gender role change as occurring in 39–64% of individuals with 17β-hydroxysteroid dehydrogenase III deficiency raised as girls.

Surgery is sometimes performed to alter the appearance of the genitals. However many surgeries performed on intersex people lack clear evidence of necessity, can be considered as mutilating, and are widely considered to be human rights violations when performed without the informed consent of the recipient.

XX males are sterile due to low or no sperm content and there is currently no treatment to address this infertility. Genital ambiguities, while not necessary to treat for medical reasons, can be treated through the use of hormonal therapy, surgery, or both. Since XX male syndrome is variable in its presentation, the specifics of treatment varies widely as well. In some cases gonadal surgery can be performed to remove partial or whole female genitalia. This may be followed by plastic and reconstructive surgery to make the individual appear more externally male. Conversely, the individual may wish to become more feminine and feminizing genitoplasty can be performed to make the ambiguous genitalia appear more female. Hormonal therapy may also aid in making an individual appear more male or female.