What is the origin of the forces generating chromosome and spindle movements in mitosis? Both microtubule dynamics and microtubule-dependent motors have been proposed as the source of these motor forces. Cytoplasmic dynein and kinesin are two soluble proteins that power membranous organelle movements on microtubules. Kinesin directs movement of organelles to the 'plus' end of microtubules, and is found at the mitotic spindle in sea urchin embryos, but not in mammalian cells. Cytoplasmic dynein translocates organelles to the 'minus' end of microtubules, and is composed of two heavy chains and several light chains. We report here that monoclonal antibodies to two of these subunits and to another polypeptide that associates with dynein localize the protein to the mitotic spindle and to the kinetochores of isolated chromosomes, suggesting that cytoplasmic dynein is important in powering movements of the spindle and chromosomes in dividing cells.