Despite the acknowledged importance of the stratum corneum in limiting water loss and in controlling skin permeability, the basis for these functions remains unknown. To pinpoint those factor(s) of importance for cutaneous barrier function, we correlated the thickness, number of cell layers, and lipid composition of leg vs. abdominal stratum corneum samples with penetration of 3H-water and 14C-salicylic acid across the same tissue sample. Viable upper epidermal sheets were obtained by incubating fresh autopsy or amputation full-thickness skin with staphylococcal exfoliatin. Each sheet was divided into 3 portions. The first piece was mounted in a diffusion cell for penetration studies. The second stratum corneum sample was frozen sectioned, stained with the fluorochrome, ANS, and measured with a micrometer eyepiece. The 3rd piece was pooled with other leg (n = 6) and abdomen (n = 15) specimens for determination of lipid weight percent. In all cases, leg stratum corneum was congruent to 2 times more permeable than abdominal stratum corneum to water and slightly more permeable to salicylic acid, as well. Penetration of both substances correlated inversely with lipid weight % of leg (mean = 3.0%) vs. abdomen (mean = 6.8%), but neither the penetration of water nor of salicylic acid was influenced by the number of cell layers or the thickness of the stratum corneum. We conclude that: differences in the thickness and the number of cell layers in the stratum corneum are insufficient to account for differences in percutaneous transport across leg and abdomen, and that total lipid concentration may be the critical factor governing skin permeability.