Abstract An equation is derived which describes the steady-state diffusion of non-electrolytes in the absence of water flow across epithelial brush borders. It is shown that the penetration rate depends on the relative values of the following quantities: the permeability constant of the cell membrane, the diffusion coefficients of the permeant inside and in the space surrounding the microvilli, the number and dimensions of the microvilli of the brush border, and the diffusional resistance of the cell body and the basal cell membrane. For substances for which the brush border rather than the basal cell membrane constitutes the main diffusional barrier numerical data were obtained which allow a conversion of empirical permeability constants as calculated under the assumption of a smooth epithelial surface into “true” permeability constants per cm 2 brush border surface. Only the latter constants are strictly comparable with those observed in other cells. The results apply to the general case where the concentration gradient across the membrane of the microvilli and hence the driving force for diffusion varies over the length of the villi.