Tremendous efforts have been made to develop better vaccines and diagnostic markers for the effective control of tuberculosis. Recently, we reported that the Triton X-100 soluble protein (TSP) of Mycobacterium tuberculosis induced strong T-cell proliferation and IFN-$gamma$ production in humans, and also conferred a significant level of protection against tuberculosis in a mouse model. In this study, the TSP was prepared by Triton X-100 extraction of Mycobacterium tuberculosis bacilli, which was followed by Triton X-114 phase partitioning. Western blot analysis using sera of 177 active pulmonary tuberculosis patients and 323 healthy individuals revealed that the TSP contained a immunodominant 40-kDa antigen specifically reacting with some sera from pulmonary tuberculosis patients. The 40-kDa antigen was purified by ion-exchange chromatography, and partially characterized by two-dimensional gel electrophoresis and N-terminal sequencing. Results of this study suggest that 40-kDa molecule of the TSP antigen from the cell surface of Mycobacterium tuberculosis can be used as a serodiagnostic marker as well as a potential vaccine candidate against tuberculosis.