The museum is situated in the old refectory of the convent erected by Augustinian hermit friars alongside the church of Santo Spirito. This large refectory, built in fourteenth-century Gothic style, is the only room in the renowned monumental complex of Santo Spirito to have maintained its original structure. Its former function is recalled by the imposing fourteenth-century fresco by Andrea Orcagna and assistants that decorates the whole eastern wall, with fragments of a Last Supper at the bottom and a superb Crucifixion with a singular descriptive style at the top. The fresco is not only one of the best works by Orcagna, but also one of the largest wall paintings from the fourteenth century to have come down to us.
Since 1946 the Cenacolo of Santo Spirito has housed the prestigious collection of sculptures, paintings, decorative artworks and antique furniture donated to the Florence City Council by collector and antiquarian Salvatore Romano (Meta di Sorrento, 1875 – Florence, 1955). Not only did he take care of personally setting up this museum and until his death acting as honorary director (a role later taken on by his son Francesco), but it was also his last wish to rest in peace in the monumental sarcophagus located against the wall in front of the fresco.
Among the most significant works are an Angel and a Caryatid (or Virtue) by Tino di Camaino, two fragments of reliefs attributed to Donatello and a Madonna and Child attributed to the circle of Jacopo della Quercia.