The original structure dates from 1100. It became the Church and Monastery of St. Mark in 1299, later passing under the protection of the Medici family in the early 15th century when Cosimo the Elder started using it regularly for his spiritual retreats.

The church had been taken over by the Dominican friars a few years earlier and Cosimo gave Michelozzo the commission of restoring it (1436-43); he also carried out the fine sacristy as well as the splendid architectural solutions used inside the monastery for the corridor of cells and the library.

Fra Angelico, a friar and artist who, like Giotto, came from the Mugello, lived at the monastery during this period (from 1435) and carried out some of his finest works here. The Florentine Humanist Academy used to meet in the "Gardens" of St. Mark, while the refectory with its Last Supper was used first by Cosimo and later by Lorenzo the Magnificent to assemble some of the finest intellects of the time.The museum occupies a vast area of the Dominican convent of San Marco and preserves much of its original atmosphere. Founded in 1436 and designed by the architect Michelozzo, the convent played an important role in the cultural and religious life of Florence, especially at the time of Savonarola.

The museum owes its renown especially to the paintings of Fra Angelico, one of the great artists of the Renaissance, who frescoed extensive parts of the convent. Other works by Fra Angelico, of various provenance, were assembled here in the 20th century, resulting in a remarkable collection of the artist's works. There is also an important collection of 16th-century paintings including numerous works by Fra Bartolomeo. The museum has a section devoted to fragments of sculpture and architecture from buildings of the city centre which were demolished in the 19th century.