Background & Aim In regenerative medicine, Wharton's jelly mesenchymal stromal cells (WJ-MSCs) have a growing interest in its properties similar to those of bone marrow cells and the immunomodulatory potential. Tissue engineering combines the design of biomaterials and the potential of cells implanted in the scaffold that interacts with the added growth factors, and its main objective is to return morphology and function by providing a biological mechanically stable niche. This evidence has directed the approach of tissue regeneration towards the use of decellularized native cell matrices. The mature organs and tissues adequately decellularized, can be used as scaffolds, because the immunogenic determinants present in the cells are eliminated, conserving a native MEC, in which its structure, architecture and mechanical properties are preserved, along with the bioavailability of proteins and growth factors necessary for tissue regeneration. We aimed is provide a temporary bone biomimetic scaffold capable of inducing osteogenic differentiation of Wharton's jelly mesenchymal stromal cells. Methods, Results & Conclusion We decellularized, demineralized, and hydrolyzed the collagen protein of the bovine bone. Then we evaluate the structure and decellularization of the scaffolds. We performed scanning electron microscopy and chemical analysis by EDS. Raman Spectroscopy and FTIR techniques determined the chemical composition by identifying functional groups and the fingerprint of type I collagen protein. We aisled WJ-MSCs and characterize surface antigens profiles of WJ-MSCs; we kept the cells in culture for 3,7,14,21 and 28 days to evaluate the biocompatibility of the scaffolds and the cellular response in vitro. On histology sections, the scaffolds showed the conservation of the structure of the ECM and cell removal. We determined that the main component is collagen type I with different degrees of denaturation depending on the concentration of hydrochloric acid (HCl) by Raman and FTIR. We eliminate the mineral phase according to the guidelines established for the FDA for demineralized scaffolds. The scaffolds also showed excellently swelling and mechanical stability. In the in vitro cell culture, the scaffolds have no cytotoxic effect and mineralized matrix deposit after 28 days in culture in the absence of growth factors or inducers of mineralization. Future studies using animal subjects are required to evaluate the degree of mineralization and scaffold response.