Enrico Castellani trained as an artist in Belgium. He attended courses in painting and sculpture at the Académie des Beaux-Arts while also studying architecture at the École Nationale Supérieure, where he took his diploma in 1956.
In 1957 Enrico Castellani returned to Italy. In Milan he came into contact with Piero Manzoni, Lucio Fontana, Vincenzo Agnetti, Agostino Bonalumi, Yves Klein and the German artists known as ZERO. The art produced by ZERO artists Heinz Mack, Otto Piene and Günther Uecker, who held the legendary evening shows of their work in Düsseldorf in 1957/58, exerted an enormous influence on the Italian avant-garde. In 1958 Enrico Castellani was committed to "Movimento Arte Nucleare", a group to which Manzoni and Fontana also belonged. That same year Enrico Castellani worked with Manzoni on the journal "Il Gesto". Enrico Castellani's first solo show of his work took place at the Galerie Kasper in Lausanne in 1959. In December 1959 Castellani and Manzoni opened Galleria Azimut in Milan. That same month the first issue of the journal "Azimuth" – a term from astronomy meaning an arc of the sky extending from the zenith to the horizon, which it cuts at right angles – was published.

The late 1950s saw Castellani's "Superficie Nera" – monochrome pictures on canvases he worked up with nails and a nail gun to give the surface a uniform relief texture that produced effects of light and shade through its indentations and protrusions. In the years that followed, Enrico Castellani continued to refine this technique and also experimented with other materials. In the 1970s and 1980s, for instance, he also used aluminum as the material for his supports. The Azimuth artists were against mimetic art; instead, they felt the object as such should be the work of art. The work possesses reality and authenticity in its own right. Light and shade, space and structure should be contained in the work itself and should not be elicited in the viewer by the use of sensory deception.

In 1964 Enrico Castellani participated in the Venice Biennale, where he was given a room of his own for his work. In 1968 he showed works at documenta 4. Numerous other exhibitions and distinctions followed that attest to Enrico Castellani's importance on an international scale. Enrico Castellani has lived in Celleno near Viterbo since 1975.