BACKGROUND A weakening of the collagen fibers of the lamina propria of the tympanic membrane is a prerequisite for the formation of a retraction pocket. The various collagen types differ in tensile strength. The distribution of the different collagen types in the lamina propria of the healthy human tympanic membrane has not been reported before. METHODS Immunohistochemical staining for collagen Types I, II, III, and IV in healthy human tympanic membranes harvested during translabyrinthine surgery for acoustic neuroma. The staining was semiquantified using light microscopy. RESULTS Collagen Type II was the most abundant collagen of the lamina propria. When subdividing the staining between the 2 fiber layers of the lamina propria, it was observed that the inner layer was enriched with Type III relative to Type II, as reflected in staining patterns. In the outer radial fiber layer of the lamina propria, staining for collagen Type II was predominant.The fibrous annulus could be subdivided into an inner and an outer portion by immunohistochemistry. The inner portion stained strongest for collagen Type II and to a lesser extent for Type I. The outer portion stained strongest for collagen Type III and Type I. CONCLUSIONS The differences in distribution of collagen types in the different fiber layers of the lamina propria suggest that the lattice of connective tissue supporting the tympanic membrane is not uniform. Understanding the differences in collagen type distribution and in the physical properties of the individual collagen types themselves may contribute to a comprehensive model of retraction pocket pathogenesis.