The intercellular lipid structure of the stratum corneum (SC) plays a key role in skin barrier function. A depth profile of the intercellular lipid conformation and the lipid lateral packing order were measured in vivo in the human SC using confocal Raman microscopy. The depth profiles of the 2880 cm(-1)/2850 cm(-1) peak ratio intensity, which represent the C-H stretching and lateral packing order of lipids, and the 1080 cm(-1)/(1130 cm(-1) + 1060 cm(-1)) peak ratio, which represents the C-C skeleton vibration and trans-gauche conformation order of lipids, were investigated. The influence of keratin on the lipid peaks at 2850 cm(-1) and 2880 cm(-1) was excluded by the developed mathematical algorithm. The results show that the trans-conformation and lateral packing order of the intercellular lipids reach their maximum value in the SC at 20-40% of its depth and then decrease towards the stratum granulosum. These results show that at a depth of 20-40% (normally corresponding to a depth of 4-8 μm) the SC exhibits the most ordered lipids and therefore the highest skin barrier function. The lateral packing of lipids is more disordered on the surface and in the deeper parts of the SC, which may be associated with a reduced skin barrier function.