The inner and outer arms of the flagellar axoneme generate the forces needed for flagellar movement; these arms contain ATPases called dyneins. To date, there has been no method for studying the mechanochemical transducing activity of isolated dyneins. Recently, it was found that the brain microtubule-associated protein (MAP) 1C is a microtubule-activated ATPase with the structural and force-producing properties of dynein. MAP 1C translocates microtubules in an in vitro gliding assay, suggesting that such an assay could also be used with axonemal dyneins. Here, we demonstrate that outer-arm dynein isolated from sea urchin (Strongylocentrotus purpuratus) sperm and adsorbed to a glass coverslip can translocate calf-brain microtubules along the surface of the coverslip. Our results conclusively demonstrate that outer-arm dynein by itself is capable of generating shearing forces. The ability to examine the force-generating properties of flagellar dynein in vitro should greatly facilitate studies of the mechanism of action of this important mechanochemical transducer.