OBJECTIVE To investigate the relationship between three purified inner ear antigens and autoimmune inner ear disease (AIED), and to determine their expressions in normal guinea pig cochleas. METHODS Guinea pigs were divided into group B, group C and group D and immunized respectively with three subcomponents of crude inner ear antigens (31,000, 42,000-45,000 and 60,000 proteins). Hearing thresholds, serum IgG levels and morphological changes of inner ear were observed. The protein expressions of these antigens were examined by immunohistochemistry. In the control group, antigens were replaced by gel homogenate of polyacrylamide. RESULTS There were no significant differences of hearing threshold among groups before immunization with inner ear antigens (F = 0.07, P > 0.05). There were no significant changes of hearing thresholds and inner ear morphology in group C and control group. Hearing thresholds dropped in part of animals immunized with 31,000 or 60,000 protein. Hearing threshold of group B and D changed significantly after immunization, as compared with the control group(F = 9.12, P < 0.01). Serum IgG levels increased significantly in all experimental groups compared with the control group(F = 7.46, P < 0.01). The 31,000 protein distributed strictly in cochlear nerve, and 42,000-45,000 or 60,000 protein distributed widely, including the spiral ganglion, Corti's organ, stria vascularis and spiral ligament. CONCLUSIONS Two subcomponents of 31,000 and 60,000 in crude inner ear antigen could induce autoimmune inner ear disease. The distribution of 31,000 protein was more tissue specific and might be used as a marker protein for clinical diagnosis of autoimmune inner ear disease.