To isolate and characterize the Beagle stem cells from apical papilla.Apical papilla was severed from the end of freshly extracted Beagle's young permanent upper anterior teeth, and digested by collagenase type I and dispase for cell culture. The isolated cells were investigated for stem cell properties by analyzing their colony-forming efficiency, growth characteristics and the expression of mesenchymal stem cell markers; and evaluating their multidifferentiation potentials including osteogenic, adipogenic, and chondrogenic potentials. Additionally, the cells were transplanted subcutaneously into immunocompromised mice to observe the mineral tissue formation.Our study showed that a clonogenic, rapidly proliferative population of cells existed in Beagle's apical papilla, and these cells had a significantly higher colony-forming rate than the stem cells from apical papilla derived from humans (P<0.001). These cells had multilineage differentiation ability including osteogenic, adipogenic and chondrogenic potentials. Mineralized nodules were formed after osteogenic induction, lipid droplets were found after adipogenic induction, and the pellets showed positive immunohistochemical staining for collagen II after chondrogenic induction. These cells also expressed the mesenchymal stem cell markers including STRO-1 and CD146, while negative for CK. Moreover, these cells transplanted with hydroxyapatite in immunocompromised mice could form mineral tissue and pulp-dentin complex-like tissue.There are stem cells from the apical papilla which have high proliferation ability and multilineage differentiation potential existing in the Beagle's apical papilla.