He served as an apprentice in the workshop of Cosimo Rosselli and then formed a workshop with the painter Mariotto Albertinelli (Albertinelli, Mariotto). His early works, such as the Annunciation (1497), were influenced by the balanced compositions of the Umbrian painter Perugino and by the sfumato (smoky effect of light and shade) of Leonardo da Vinci. Influenced by the preaching of the Florentine Dominican religious reformer Girolamo Savonarola (Savonarola, Girolamo), Bartolommeo joined the Dominican order in 1500 and gave up painting. He began painting again in 1504, producing devotional paintings mostly at the service of his order. His Vision of St. Bernard (completed 1507) shows him achieving the transition from the subtle grace of late Quattrocento painting to the monumentality of the High Renaissance style.

Bartolommeo visited Rome in 1514, where he saw Raphael's mature work and Michelangelo's frescoes on the ceiling of the Sistine Chapel. In response Bartolommeo's art took on a greater power of dramatic expression, as in the Madonna della Misericordia (1515) and the Pietà (c. 1515). Despite Bartolommeo's assimilation of the progressive currents of his time, his art is restrained, conservative, and somewhat severe, and he painted religious subjects almost exclusively. His production of drawings and preparatory sketches shows a delicate sensitivity and technical superiority. His landscapes are among the most notable of his time.