The subunit composition and dimeric species of tropomyosin with respect to its alpha and beta subunits have been analyzed from several physiologically different types of skeletal muscle of the rabbit using one- and two-dimensional gel electrophoresis. The findings indicate that there are two types of tropomyosin distributions in rabbit muscles: one in which alpha 2 is the prevalent species and another in which alpha beta is prevalent. While the alpha 2-prevalent pattern is characteristic of some fast muscles, notably longissimus dorsi and psoas, it is not the only pattern found in fast muscles. Plantaris, which is histochemically indistinguishable from longissimus dorsi, exhibits an alpha beta-prevalent distribution. This observation shows that there are at least two types of fast white muscle fibers which differ in their tropomyosin composition and species. The alpha beta-pattern is also found in fast red, slow, and mixed muscles. The fast and slow alpha beta-prevalent patterns can be distinguished electrophoretically because of heterogeneity of their subunits. Finally, the relative abundance of the different forms of tropomyosin in each of the muscles studied indicates that the tropomyosin dimer does not assemble randomly from alpha and beta subunits, but that alpha beta is assembled preferentially in vivo.