Work in mouse has implicated cilia motility and leftward nodal flow as the mechanism for breaking left-right symmetry. In zebrafish, it is assumed that Kupffer's vesicle is analogous to the mouse node. However, its architecture is different and the fluid dynamics inside Kupffer's vesicle is not completely understood. We show that cells lining both the dorsal roof and the ventral floor of Kupffer's vesicle possess posteriorly pointed cilia that rotate clockwise when viewed apically. Analysis of bead movements within Kupffer's vesicle shows a net circular flow but the local flow differs in direction depending on the location within the vesicle. Histological analysis suggests that the orientation of the cells at anterior–dorsal region likely direct net flow in the vesicle. Our data suggest that the plane of the circular net flow is tilted with respect to the D–V axis, which may be converted to a local leftward flow in the anterior–dorsal region of the vesicle. Developmental Dynamics 237:3602–3612, 2008. © 2008 Wiley-Liss, Inc.