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To compare morphologic features of keratoconus as observed in vivo with a slit scanning confocal microscope and in vitro using light microscopy.Slit scanning confocal microscopy (CM) was used to evaluate the central cornea of 29 keratoconic subjects (mean age, 31 ± 10 years; range, 16-49). Light microscopy (LM) examination was performed on 2 of the keratoconic corneas post-keratoplasty.With CM, the epithelium appeared more abnormal with increasing severity of keratoconus. In severe disease, the epithelium displayed the following characteristics: superficial cells were elongated and spindle shaped, wing cell nuclei were larger and more irregularly spaced, and basal cells were flattened. These findings were confirmed by LM. Images obtained using CM revealed disruption to Bowman's layer and the occasional presence of epithelial cells and stromal keratocytes. This was shown with LM to be due to breaks in Bowman's layer. Stromal haze and hyperreflectivity observed with CM corresponded with apical scarring seen on slit-lamp biomicroscopy. Hyperreflective keratocyte nuclei observed with CM are thought to indicate the presence of fibroblastic cells seen with LM. Increasing levels of haze detected with CM were found with LM to be due to fibroblast accumulation and irregular collagen fibers. Descemet's membrane appeared normal with both CM and LM. Evidence of endothelial cell elongation was apparent in 1 subject with CM.The current study confirms the application of CM for assessing morphologic alterations to the epithelium, Bowman's layer, and stroma in keratoconus. Many of the tissue changes observed with CM could be reconciled with observations made using LM. This work provides a framework against which tissue changes in keratoconus can be studied in a clinical context in vivo using CM.