Differential effects of tissue processing on human embryonic and fetal skin

To most accurately evaluate quantitative data from studies of developing epidermis, the effects of tissue processing on human embryonic and fetal skin (8-20 weeks gestational age) were examined using two different techniques: 1) EDTA-separated epidermal sheets that were briefly fixed in 2% paraformaldehyde, processed through Permount infiltration, and prepared as whole mounts on glass slides, and 2) skin that was fixed in Karnovsky's fixative and embedded in Epon. Based on en face measurements of surface area before and after tissue processing, both procedures caused differential, age-dependent shrinkage. However, the trend of increasing shrinkage was inversely related to increasing age in the paraformaldehyde-fixed epidermal sheets (y = 57.14 + 1.26x, where x = gestational age in weeks and y = % of original surface area), but directly correlated with aging in the Karnovsky-fixed skin (y = 955.62 - 232.77x + 20.38x2). Shrinkage of epidermal sheets occurred during the dehydration and clearing steps, whereas most of the dimensional changes in whole skin took place during fixation in Karnovsky's. These differences are probably due to greater cross-linking of proteins and longer fixation time in the more concentrated and fast-acting Karnovsky's, as well as the influence of increasing quantities of fibrous proteins in the dermis of whole skin.