The embryonic precursors of the avian slow (type I and III) and fast (type II) fibers can be distinguished from each other early in muscle formation (stage 28, V. Hamburger and H. L. Hamilton, J. Morphol, 88, 49-92, 1951) on the basis of the differential sensitivity of their myosin ATPases. To test the neural dependence of fiber type differentiation, the source of motor innervation was eliminated by excision of the brachial neural tube at stages 16-18 before muscles are innervated. Removal of the brachial neural tube did not affect the number of primary myotubes in a sample muscle of the forelimb (ulnimetacarpalis dorsalis, UMD) up until stage 36. Myosin ATPase staining at a variety of pHs revealed the typical patterns of fiber types in muscles of neural-tube free embryos in stages 35-37. These muscles included the anterior latissimus dorsi, brachialis, and UMD which showed presumptive type III staining (type IIIEMB), the pronator superficialis and flexor carpi ulnaris which showed embryonic type II staining (type IIEMB), and the triceps brachii muscles which showed characteristic arrangements of both type IEMB and type IIEMB fibers. The normal patterns of type IEMB and type IIEMB myotubes were also seen in muscles containing a heterogeneous mixture of fiber types such as the biceps brachii, extensor metacarpi radialis, and adductor indicis muscles, although the intensity of acid-stable ATPase staining of the type IEMB myotubes in these muscles was lower than in innervated muscles. It is concluded that the earliest differentiation of muscle fiber types is independent of the nervous system.