Dental pulp stem cells were isolated from natal teeth of a healthy newborn female. Cells adhered to the culture plate and exhibited a large, flattened or fibroblast-like morphology. Cells expressed mesenchymal marker and had the capacity to differentiate into mature adipocytes, myoblasts, neuroglial cells, chondroblasts and osteoblasts in vitro.
The pulp tissue was digested, using collagenase type I, to form single-cell suspensions. The cells were then cultured in MEM-Earle supplemented with fetal bovine serum (FBS; 15%) and 100 IU/ml penicilin and 100 μg/ml streptomycin. Cells from one tooth were seeded into two 25 cm2 plastic tissue culture flasks and incubated for 3 days. On the third day, red blood cells and other nonadherent cells were removed and the medium was replaced. Adherent cells were cultured until they reached 70% confluency and were defined as passage
zero (P0) cells.