Anaïs Nin was born in Neuilly, France, to two artistic parents. Her father, Joaquín Nin, was a Cuban pianist and composer, and her mother Rosa Culmell was a classically trained Cuban singer of French and Danish ancestry. Her paternal great-grandfather had fled France during the Revolution, going first to Saint-Domingue, then New Orleans, and finally to Cuba where he helped build that country's first railway. After her parents separated, her mother moved Anaïs and her two brothers, Thorvald Nin and Joaquin Nin-Culmell, from Barcelona to New York City. According to her diaries, Volume One, 1931–1934, Nin abandoned formal schooling at the age of 16 and began working as a model.

On March 3, 1923, in Havana, Cuba, Nin married her first husband, Hugh Parker Guiler (1898—1985), a banker and artist, later known as "Ian Hugo" when he became a filmmaker of experimental films in the late 1940s. The couple moved to Paris the following year, where Guiler pursued his banking career and Nin began to pursue her interest in writing; in her diaries she also mentions having trained as a flamenco dancer in Paris in the mid-to-late 1920s. Her first published work was a critical evaluation of D. H. Lawrence called An Unprofessional Study. She also explored the field of psychotherapy, studying under the likes of Otto Rank, a disciple of Sigmund Freud.

Nin left Paris in the late summer of 1939, when residents from overseas were urged to leave France due to the upcoming war and returned to New York City with Guiler (who was, on his own wish, all but edited out of her published diaries and whose role in her life is therefore difficult to gauge). During the war, Nin sent her books to Frances Steloff of the Gotham Book Mart in New York for safekeeping.

According to her diaries,Vol.1, 1931—1934, Nin shared a bohemian lifestyle with Henry Miller during her time in Paris. Her husband Guiler is not mentioned anywhere in the published edition of the 1930s parts of her diary (Vol.1—2) although the opening of Vol.1 makes it clear that she is married. Nin appeared in the Kenneth Anger film Inauguration of the Pleasure Dome (1954) as Astarte; in the Maya Deren film Ritual in Transfigured Time (1946); and in Bells of Atlantis (1952), a film directed by Guiler under the name "Ian Hugo" with a soundtrack of electronic music by Louis and Bebe Barron.

In 1947, at the age of 44, she met and began living with Rupert Pole (1919—2006), sixteen years her junior. On March 17, 1955, she married him at Quartzsite, Arizona, returning with Pole to live in California. Guiler remained in New York City and was unaware of Nin's second marriage until after her death in 1977.

After Guiler's death in 1985, the unexpurgated versions of her journals were commissioned by Pole.

Nin often cited authors Djuna Barnes and D. H. Lawrence as inspirations. She states in Volume One of her diaries that she drew inspiration from Marcel Proust, André Gide, Jean Cocteau, Paul Valéry, and Arthur Rimbaud.

Nin once worked at Lawrence R. Maxwell Books located at 45 Christopher Street.

Anaïs Nin is perhaps best remembered as a diarist. Her journals, which span several decades, provide a deeply explorative insight into her personal life and relationships. Nin was acquainted, often quite intimately, with a number of prominent authors, artists, psychoanalysts, and other figures, and wrote of them often, especially Otto Rank. Moreover, as a female author describing a primarily masculine constellation of celebrities, Nin's journals have acquired importance as a counterbalancing perspective.

Previously unpublished works are coming to light in A Café in Space, the Anais Nin Literary Journal, which most recently includes "Anais Nin and Joaquín Nin y Castellanos: Prelude to a Symphony...Letters between a father and daughter."

Nin is hailed by many critics as one of the finest writers of female erotica. She was one of the first women to explore fully the realm of erotic writing, and certainly the first prominent woman in modern Europe to write erotica. Before her, erotica written by women was rare, with a few notable exceptions, such as the work of Kate Chopin.

According to Volume I of her diaries, 1931—1934, published in 1966 (Stuhlmann), Nin first came across erotica when she returned to Paris with her mother and two brothers in her late teens. They rented the apartment of an American man who was away for the summer, and Nin came across a number of French paperbacks: "One by one, I read these books, which were completely new to me. I had never read erotic literature in America They overwhelmed me. I was innocent before I read them, but by the time I had read them all, there was nothing I did not know about sexual exploits I had my degree in erotic lore."

Faced with a desperate need for money, Nin, Miller and some of their friends began in the 1940s to write erotic and pornographic narratives for an anonymous "collector" for a dollar a page, somewhat as a joke. (It is not clear whether Miller actually wrote these stories or merely allowed his name to be used.) Nin considered the characters in her erotica to be extreme caricatures and never intended the work to be published, but changed her mind in the early 1970s and allowed them to be published as Delta of Venus and Little Birds.

Nin was a friend, and in some cases lover, of many leading literary figures, including Henry Miller, Antonin Artaud, Edmund Wilson, Gore Vidal, James Agee, James Leo Herlihy, and Lawrence Durrell. Her passionate love affair and friendship with Miller strongly influenced her both as a woman and an author. The rumor that Nin was bisexual was given added circulation by the Philip Kaufman film Henry & June. This rumor is dashed by at least two encounters Nin writes about in her third unexpurgated journal, Fire. The first is with a patient of Nin's (Nin was working as a psychoanalyst in New York at the time), Thurema Sokol, with whom nothing physical occurs. She also describes a ménage à trois in a hotel, and while Nin is attracted to the other woman, she does not respond completely (229—31). Nin confirms that she is not bisexual in her unpublished 1940 diary when she states that although she could be attracted erotically to some women, the sexual act itself made her uncomfortable. What is irrefutable is her sexual attraction to men.

Nin's first unexpurgated journal, Henry and June, makes it clear, despite the notion to the contrary, that she did not have sexual relations with Miller's wife, June. While Nin was stirred by June to the point where she says (paraphrasing), "I have become June," she did not consummate her erotic feelings for her. Still, to both Anais and Henry, June was a femme fatale...irresistible, cunning, erotic. Nin gave June money, jewelry, clothes, oftentimes leaving herself broke. In her second unexpurgated journal, Incest, she wrote that she had an incestuous relationship with her father, which was graphically described (207—15). When Nin's father learned of the title of her first book of fiction, House of Incest, he feared that the true nature of their relationship would be revealed, when, in fact, it was heavily veiled in Nin's text.

In 1973 Anaïs Nin received an honorary doctorate from the Philadelphia College of Art. She was elected to the United States National Institute of Arts and Letters in 1974. She died in Los Angeles, California on January 14, 1977; her body was cremated, and her ashes were scattered over Santa Monica Bay. Rupert Pole was named Nin's literary executor, and he arranged to have new unexpurgated editions of Nin's books and diaries published between 1985 and his death in 2006.

Philip Kaufman directed the 1990 film Henry & June based on Nin's novel From the Unexpurgated Diary of Anaïs Nin. She was portrayed in the film by Maria de Medeiros.