Biological sex: a complex reality

When we talk about biological sex, we refer to a set of physical characteristics of the human body that are traditionally used to classify people as male or female. It is one of the most fundamental concepts in biology, yet also one of the most misunderstood in public debate. In this article, we explore what biological sex really is, how it is determined, what its components are, and why the scientific reality is more nuanced than commonly thought.

What biological sex means

The National Institutes of Health (NIH) of the United States defines biological sex as a “multidimensional biological construct based on anatomy, physiology, genetics, and hormones” [13]. This definition is important because it underscores an often overlooked aspect: biological sex is not a single parameter, but the result of the interaction of several biological components that, in most cases, align consistently, but that can also follow independent paths.

The main components of biological sex are:

• Chromosomal sex: the sex chromosome complement in cells
• Gonadal sex: the type of gonads present (ovaries or testes)
• Hormonal sex: the levels and types of sex hormones produced and the sensitivity of receptors to those hormones
• Phenotypic or anatomical sex: the external and internal sexual characteristics, including the genitals and reproductive system

Each of these dimensions contributes to defining a person’s biological sex, and each deserves to be understood in detail.

Chromosomal sex

Sex chromosomes: XX and XY

In humans, cells typically contain 23 pairs of chromosomes, for a total of 46. The last pair is the sex chromosomes: in the most common configuration, females have two X chromosomes (karyotype 46,XX) and males have one X chromosome and one Y chromosome (karyotype 46,XY). This is the most basic level of sex determination [9].

The Y chromosome carries a fundamental gene called SRY (Sex-determining Region Y), which encodes a protein known as the testis-determining factor [4]. This gene acts as a biological switch: its activation, around the sixth to seventh week of embryonic development, initiates a cascade of molecular events that will lead to the development of testes [4]. Without the activation of the SRY gene, the gonads develop as ovaries.

However, the role of the SRY gene, while central, is not the only factor at play. As MedlinePlus explains, the SRY gene works by activating another gene, SOX9, which in turn directs the differentiation of Sertoli cells and the formation of testicular tissue [4]. It is a complex chain, and a mutation at any point in this chain can alter the course of sexual development.

Chromosomal variations

Not all people have an XX or XY karyotype. There are numerous natural chromosomal variations that demonstrate how chromosomal sex is not always reducible to only two possibilities [8].

Klinefelter syndrome (47,XXY): occurs when an individual is born with two X chromosomes and one Y chromosome. It is one of the most common chromosomal variations, with an estimated frequency of about 1 in 500-600 male births [7]. People with this condition are generally assigned male at birth, but may present characteristics such as tall stature, gynecomastia (breast tissue development), reduced testosterone production, and, often, infertility [7]. Many people with Klinefelter syndrome never receive a diagnosis, because symptoms can be mild.

Turner syndrome (45,X0): occurs when a person has only one X chromosome, without the second sex chromosome. The frequency is about 1 in 2,000 female births [12]. People with this condition are generally assigned female at birth, but present characteristics such as short stature, absence of spontaneous pubertal development, and infertility [12].

There are also other variations, such as the 47,XYY karyotype, the 47,XXX karyotype (trisomy X), and chromosomal mosaicisms, in which different cells of the same organism have different chromosome complements — for example, some cells XX and others XY. These variations demonstrate that chromosomal sex is not a perfectly binary system, but presents a variety of natural configurations [8].

A particularly interesting case is that of XY females: people with a 46,XY karyotype who, due to mutations in the SRY gene or other conditions, develop a completely female phenotype (Swyer syndrome). Similarly, there are XX males: people with a 46,XX karyotype in which a portion of the Y chromosome containing the SRY gene has translocated onto an X chromosome, leading to the development of male characteristics [4].

Gonadal sex

Gonads — ovaries and testes — are the organs that produce reproductive cells (eggs and sperm) and sex hormones. In most cases, gonadal sex corresponds to chromosomal sex: XX individuals develop ovaries and XY individuals develop testes.

However, as we have seen, this is not always the case. Some people may have gonadal tissue that does not fully differentiate as ovarian or testicular, or may present ovotestes — gonads that contain both ovarian and testicular tissue. This condition, once called “true hermaphroditism” and now defined as ovotesticular DSD (Difference of Sex Development), is rare but documented in the medical literature.

Gonadal sex is important because the gonads are the main source of sex hormones that shape the body during prenatal development, puberty, and throughout adult life. But the type of gonad present does not automatically determine the hormonal profile: ovaries also produce testosterone (in smaller amounts) and testes also produce estrogens.

Hormonal sex

Sex hormones — particularly testosterone, estrogens, and progesterone — play a fundamental role in the development of the body’s sexual characteristics. All humans produce both testosterone and estrogens, but in generally different proportions: people with testes tend to produce higher levels of testosterone, while people with ovaries tend to produce higher levels of estrogens.

However, hormonal sex depends not only on the quantity of hormones produced, but also on the sensitivity of receptors to these hormones. This aspect is crucial for understanding some conditions that challenge the binary model.

Androgen insensitivity syndrome

Androgen insensitivity syndrome (AIS) is an emblematic example of how hormonal sex can diverge from chromosomal and gonadal sex. People with complete AIS have a 46,XY karyotype and functioning testes that produce testosterone in normal or even above-average amounts. However, their cells are unable to respond to testosterone due to a mutation in the androgen receptor gene.

The result is that these individuals develop a completely female phenotype: female external genitalia, breast development during puberty, absence of typically male body hair. Often the condition is discovered only in adolescence, when menarche does not occur, or in adulthood during investigations for infertility.

Congenital adrenal hyperplasia

Congenital adrenal hyperplasia (CAH) is a genetic condition in which the adrenal glands produce excessive amounts of androgens. In people with a 46,XX karyotype, this prenatal exposure to elevated androgen levels can lead to the development of external genitalia with atypical characteristics, sometimes classified as ambiguous at birth.

5-alpha-reductase deficiency

Another significant condition is 5-alpha-reductase deficiency, an enzyme that converts testosterone to dihydrotestosterone (DHT), necessary for the development of male external genitalia during prenatal life. People with a 46,XY karyotype and this condition are often born with female-appearing or ambiguous genitalia and are assigned female at birth. With puberty, the increase in testosterone causes significant virilization: the voice deepens, muscle mass develops, and, in some cases, the genitalia change in a male direction.

These conditions demonstrate that hormonal sex is a complex system in which not only hormone levels matter, but also the body’s ability to respond to them.

Phenotypic and anatomical sex

Phenotypic sex refers to the visible and measurable sexual characteristics of the body: external genitalia, internal genitalia (uterus, prostate, vas deferens), secondary sexual characteristics that develop during puberty (breasts, body fat distribution, body hair, voice pitch, shoulder and hip width), and other physical characteristics.

In the vast majority of people, the sexual phenotype is consistent with chromosomal, gonadal, and hormonal sex. However, the conditions described in the preceding paragraphs show how the phenotype can diverge from these other components. A person with XY chromosomes can have a completely female phenotype (as in the case of complete AIS), and a person with XX chromosomes can present virilized sexual characteristics (as in the case of CAH).

It is important to emphasize that sex assigned at birth is based almost exclusively on the observation of the external genital phenotype. Routine chromosomal or hormonal tests are not performed: medical staff observe the newborn’s genitalia and assign sex accordingly. This means that the sex recorded at birth is a classification based on only one of the several components of biological sex.

Intersex conditions

The term intersex (or the more recent expressions “variations of sex characteristics” and “differences of sex development,” DSD) refers to a broad spectrum of natural conditions in which a person’s sexual characteristics do not fit within typical definitions of male or female. These variations may involve chromosomes, gonads, hormones, or anatomy, or a combination of these elements.

How common are intersex variations?

The answer to this question depends significantly on the definition used. Biologist Anne Fausto-Sterling estimated that about 1.7% of the population presents some form of variation of sex characteristics — a frequency comparable to that of red hair [6]. This estimate includes conditions such as Klinefelter syndrome, Turner syndrome, and milder forms of congenital adrenal hyperplasia.

A more restrictive estimate, proposed by researcher Leonard Sax, focuses exclusively on cases where the sexual phenotype is visibly ambiguous or where chromosomal sex is clearly incongruent with phenotypic sex, arriving at a prevalence of about 0.018% [5]. The difference between these estimates reflects a fundamental disagreement about what it means to be intersex and which conditions should fall within this category.

The OHCHR (Office of the United Nations High Commissioner for Human Rights) uses the broader estimate, stating that intersex people represent up to 1.7% of the population [11]. Regardless of the chosen estimate, intersex variations are a documented biological reality and cannot be ignored in any serious discussion of biological sex.

Biological sex as a spectrum: what recent science says

In 2015, a groundbreaking article published in Nature by science journalist Claire Ainsworth, titled Sex Redefined, brought to the general public a message that the scientific community had been discussing for some time: the idea that only two sexes exist is an oversimplification [1]. The article illustrated how the various components of biological sex do not always align consistently and how the boundaries between “male” and “female” are less clear-cut than commonly assumed [1].

In 2025, the journal Science published a review of the book Sex Is a Spectrum: The Biological Limits of the Binary by anthropologist Agustin Fuentes, a professor at Princeton [2]. The book traces the origin and evolution of sex in the animal kingdom, describes the variety of ways organisms are female, male, or both, and presents fossil and archaeological evidence of diversity in sexual structures, gender roles, and family configurations throughout human history [3].

As Fuentes himself clarified: “I’m not saying there are more than two sexes — there are male and female. But male and female are not essential entities that are distinct from one another” [3]. In other words, the classification into two categories is useful and functional in most cases, but it does not capture the entire range of biological variability.

A fundamental aspect is that this complexity does not concern only extreme cases or rare conditions. Even among people who clearly fall within the categories of male or female, there is wide variability in hormone levels, bone density, body fat distribution, muscle mass, and many other characteristics we consider “sexual.” Two men can have very different testosterone levels, and both fall within the normal range. Two women can have significantly different hormonal profiles, and both are biologically female. Biological sex, in summary, describes a bimodal distribution — with two distinct peaks — rather than an absolute dichotomy.

The difference between biological sex and gender identity

An essential aspect, often a source of confusion in public debate, is the distinction between biological sex and gender identity. These are two different dimensions of human experience, and confusing them leads to significant misunderstandings.

Biological sex refers, as we have seen, to the physical characteristics of the body: chromosomes, gonads, hormones, and anatomy. It is a biological construct that is assessed (in a simplified way) at birth through observation of the genitalia.

Gender identity, according to the American Psychological Association’s definition, is the intimate and deep sense a person has of their own gender — feeling like a man, a woman, a combination of both, neither, or something else. Gender identity is a subjective and inner experience that may or may not correspond to the sex assigned at birth.

The Canadian Institutes of Health Research (CIHR) clearly distinguish the two dimensions: sex refers to a set of biological attributes in humans and animals, while gender refers to the socially constructed roles, behaviors, expressions, and identities of girls, women, boys, men, and gender-diverse people [10].

This distinction is supported by scientific research. Neuroimaging studies have shown that the brains of transgender people present certain structural and functional characteristics more similar to those of cisgender people with the same gender identity than to those of people with the same biological sex. Twin studies have also found a significant heritable component of gender identity, with an estimated heritability index between 0.30 and 0.57. These data indicate that gender identity has its own biological foundations, distinct from biological sex in the strict sense.

Why “it’s basic biology” is an oversimplification

One of the most frequent phrases in discussions about sex and gender is: “it’s basic biology.” Those who use it generally mean to assert that there are two sexes, male and female, that these are determined by chromosomes, and that there is nothing more to say. But it is precisely biology — the advanced kind, not the basic kind — that shows us a much more detailed picture.

“Basic biology” — the kind taught in elementary and middle school — is a necessary and pedagogically useful simplification. However, as one advances in the study of biology, a complexity emerges that this simplification cannot capture:

• Chromosomes alone do not determine sex: it is the SRY gene, and the molecular cascade that follows, that guides sexual development, and this cascade can be disrupted at many points [4].
• Gonads do not always develop consistently with the chromosomal karyotype.
• Hormones act only if receptors are functional, and hormonal sensitivity varies among individuals.
• Phenotype can diverge from genotype in clinically significant ways.
• There are people with XY chromosomes and a completely female body, and people with XX chromosomes and male sexual characteristics.

Asserting that “sex is simple” is not a scientific position: it is a position that ignores decades of research in genetics, endocrinology, developmental biology, and medicine. As the NIH effectively summarizes, sex is a “multidimensional biological construct,” not a binary switch [13].

This does not mean that the classification into males and females is useless or lacking biological foundation. For the vast majority of people, the various components of sex align consistently and the binary classification is descriptive and functional. But recognizing biological complexity is essential for understanding the experience of intersex people, for informing health policies, and for having a public debate grounded in scientific evidence rather than oversimplifications.

In summary

Biological sex is a complex system composed of multiple dimensions — chromosomes, gonads, hormones, anatomy — that in most cases align consistently toward one of the two poles, male or female. However, biology naturally produces a variety of combinations that do not fit into a perfect dichotomy [1][3]. Intersex conditions, chromosomal variations, and differences in hormonal response are part of normal human biological variability [11].

Understanding this complexity does not mean denying the existence of sexual differences, but recognizing that biological reality is more nuanced than a purely binary model can capture. And understanding biological sex in its entirety is the first step toward addressing with scientific rigor the discussions about gender identity, the rights of intersex people, and health policies.