In spite of long term clinical use of decellularized bladder submucosa matrix (BSM), little is known about the active factors within this material. In this paper, we analyzed the biological factors from the decellularized BSM using ELISA, Western blotting, and immunohistochemistry for the purpose of effective utilization of this material in the field of regenerative medicine. At least 10 growth factors, including VEGF, BMP4, PDGF-BB, KGF, TGFbeta1, IGF, bFGF, EGF and TGFalpha were found to be preserved in the decellularized BSM. The existence of collagen (type 1, 2, 3, 4), laminin and elastin within the matrix was also demonstrated. The soluble BSM extracts showed a conspicuous effect on cell proliferation when added as a supplement in vitro. These findings demonstrate that growth factors and extracellular matrix in the BSM maintain valuable biological activity even after the decellularization and extraction processes, thus supporting the wide applicability of BSM in tissue regeneration. The identification and characterization of growth factors and extracellular matrix in the BSM is a prerequisite for understanding tissue regeneration using this scaffold.