Owls are known for their nocturnal hunting capability. Many owl species are able to localize prey in complete darkness just by hearing. Sound localization of strictly nocturnal owls is improved by asymmetrically arranged outer ears. According to Norberg (1977), who worked with adult owls, asymmetrical ears evolved at least four times independently among owls. What is unknown so far is how the ear asymmetry develops in the embryo. Here we examine the embryonic development of ear asymmetry in the American barn owl (Tyto furcata pratincola) in the frame of the 42 stages suggested by Köppl et al. (2005). In this species, the left ear opening in the skin is located higher than its counterpart on the right. Micro-CT scans show that in an anatomically defined coordinate system, the ear openings initially appear symmetrically as does the skull as well as the eyes, the nasal openings, the stapes and the squamosum. The ear openings are initially located ventrally in the skull. Soon after their appearance, the ear openings start to move dorso-occipitally. At the developmental stages 36–39, the left ear opening moves faster than the right one. In this way, an ear asymmetry develops within a few developmental stages. The skull and the other anatomical markers remain symmetrical. Thus, these data show that the soft tissue asymmetry in the barn owl develops already before hatching. It will be interesting to compare the time course described here with the time course of development in owl species with bony ear asymmetries.