PurposeDeposition of abnormal sub-epithelial matrix and posterior collagenous layer by epithelium and endothelium, respectively, in Fuchs' dystrophy gives us the opportunity to determine if these tissues synthesizeβig-h3.MethodsImmunohisto-/immunocytochemistry of corneas were conducted with rabbit anti-humanβig-h3 and monoclonal anti-human type VI collagen. Labeled sense and anti-senseβig-h3 oligonucleotide probes were used forin situhybridization.Resultsβig-h3-specific fluorescence was found just beneath detached epithelium in the sub-epithelial matrix, abnormal Descemet's membrane and posterior collagenous layer. Type VI collagen co-localized withβig-h3 within abnormal sub-epithelial matrix and corneal stroma adjacent to Descemet's membrane.βig-h3 mRNA was detected in corneal epithelium of dystrophic corneas.ConclusionsExpression ofβig-h3 in sub-epithelial matrix and posterior collagenous layer of Fuchs' dystrophy is consistent with the synthesis of new extracellular matrices by epithelial and endothelial tissues.βig-h3 mRNA in corneal epithelium further supports an epithelial source of this protein. Endothelial synthesis ofβig-h3 is based on circumstantial evidence due to cell loss during surgical and histological procedures. Co-localization ofβig-h3 with type VI collagen in abnormal sub-epithelial matrix and at the stromal/Descemet's membrane interface suggest this collagen in association withβig-h3 interacts with these tissues and anchors them to the adjacent stroma. Curr. Eye Res. 15: 965–972, 1996.