He was born at Męto (Mauth) near Rokycany, Bohemia. His father was a cultured man, and his mother was the sister of Raphael Georg Kiesewetter (1773-1850), the musical archaeologist and collector. Ambros was well-educated in music and the arts, which were his abiding passion. He was, however, destined for the law and an official career in the Austrian civil service, and he occupied various important posts under the ministry of justice, music being an avocation.
From 1850 onwards he became well-known as a critic and essay-writer, and in 1860 he began working on his magnum opus, his History of Music, which was published at intervals from 1864 in five volumes, the last two (1878, 1882) being edited and completed by Otto Kade and Langhaus.
Ambros became professor of the history of music at Prague in 1869. He was an excellent pianist, and the author of numerous compositions somewhat reminiscent of Felix Mendelssohn.
Ambros died at Vienna, Austria at the age of 59.