Niki was born Catherine Marie-Agnès de Saint Phalle on October 29, 1930 in France. Her father was French, her mother American. After being wiped out financially during the Great Depression, the family moved from France to the United States in 1933. She spended most of her childhood and adolescence in New York City, though strong ties were maintained with the family in France through frequent visits.

Niki enrolled at the prestigious Brearley School in New York City, but she was dismissed for painting fig leaves red on the school's statuary. She transfered to a new school shortly thereafter.

During her teenaged years, she was a fashion model; at the age of sixteen, she appeared on the cover of Life magazine (September 26, 1949), and, three years later, on the November 1952 cover of French Vogue. At 18, she eloped with childhood friend Harry Mathews and moved to Cambridge, Massachusetts. While her husband studied music at Harvard University, de Saint Phalle began to paint, experimenting with different media and styles.

In 1952, Niki moved to Paris to study theater and acting. In 1953, hospitalized for a nervous breakdown, Niki finded that painting helps her to overcome this crisis and decided to give up acting and become an artist. In Spain, Niki discovered the work of Antonio Gaudí and was deeply affected, especially by Park Güell in Barcelona, which planted the idea to create her own sculpture garden and inspired her to use diverse materials and found objects as essential elements in her art. In Paris Niki met Jean Tinguely, who will become an artistic collaborator. She is further inspired by the art of Paul Klee, Henri Matisse, Pablo Picasso, and Henri Rousseau.

In 1960, Niki and Harry separated. She set up a studio and continues her artistic experiments. She was included in an important group exhibition at the Musée d'Art Moderne de la Ville de Paris. By the end of the year Niki and Jean Tinguely moved in together, sharing the same studio and living in an artists' colony. During the 1960s, she became friends with American artists in Paris, such as Robert Rauschenberg, Jasper Johns, and Larry Rivers and his wife Clarice, with whom de Saint Phalle collaborated over the years.

In 1961, she became known around the world for her Shooting paintings. A shooting painting consisted of a wooden base board on which containers of paint were laid, then covered with plaster. The painting was then raised and de Saint Phalle would shoot at it with a .22 caliber rifle. The bullets penetrated paint containers, which spilled their contents over the painting. This "painting style" was completely new, and she travelled around the world performing shooting sessions in Paris, Sweden, Malibu, California, and the Stedelijk Museum in Amsterdam. Saint Phalle had stopped making these shooting pictures in 1963 as in her own words, ‘I had become addicted to shooting, like one becomes addicted to a drug'.

Inspired by the pregnancy of Larry Rivers' wife Clarice in 1965, Niki maked her first Nanas, archetypal female figures which are updated versions of "Every(wo)man." (The word "nana" is French for "dame" or "chick.") For the first exhibit of Nanas, Niki's first artist book was published. This developed into another of Niki's prolific art forms: hand-lettered graphic works in the form of invitations, posters, books, and other writings. During the 1960s, she also designed decors and costumes for two theatrical productions: a ballet by Roland Petit, and an adaptation of the Aristophanes play "Lysistrata."
In 1966, Niki collaborated on Hon (Swedish for "she") for the Moderna Museet, Stockholm. The outer form of Hon is a building-size giant reclining Nana with an interior environment entered from between her legs. This piece garnered worldwide attention and intensified her desire to build her own sculpture garden.

Influenced by Gaudí´s Parc Güell in Barcelona, and the garden in Bomarzo, de Saint Phalle decided that she wanted to make something similar; a monumental sculpture park created by a woman. In 1979, she acquired some land in Garavicchio, Tuscany, about 100 km north-west of Rome along the coast. The garden, called Giardino dei Tarocchi in Italian, contains sculptures of the symbols found on Tarot cards. The garden took many years, and a considerable sum of money, to complete. It opened in 1998, after more than 20 years of work.

Niki de Saint-Phalle died on May 21, 2002.