Seven galleries display selections from the permanent collection of over 17,000 works of art. Special exhibitions drawn from the collection for touring from other museums are displayed in four additional galleries.

Because the Museum serves as a resource for the teaching and study of art history, fine arts, and the humanities, the collection is comprehensive in nature. It spans the history of European, North American, and East Asian art. Areas of special strength include medieval art; European and American paintings, sculpture, and prints; photography; Japanese Edo-period painting and prints; and 20th-century Chinese painting.

More than half of the Western paintings and sculpture are on permanent exhibit. The much more numerous Western and Asian prints, drawings, photographs, and decorative arts, including the Spencer Museum's renowned quilt collection, are shown on a rotating basis in short-term, thematic exhibitions. Those objects not on view are available for scholarly examination by appointment.