The study characterises canine adipose-derived stem cells (cASCs) in comparison to human ASCs (hASCs) and tests their safety in a canine model after intravenous administration. cASCs from two dogs were cultured under hypoxic conditions in a medium supplemented with autologous serum. They were plastic adherent, spindle-shaped cells that expressed CD73, CD90, and CD44 but lacked CD45, CD14, HLA-DR, and CD34. cASCs differentiated toward adipogenic, osteogenic, and chondrogenic lineages, although adipogenic differentiation capacity was low. Blast transformation reaction demonstrated that these cells significantly suppress T-cell proliferation, and this ability is dose-dependent. Intravenous administration of a cell freezing medium, therapeutic dose of cASCs (2 × 106 live cells/kg), and five times higher dose of cASCs showed no significant side effects in two dogs. Microscopic tissue lesions were limited to only mild, non-specific changes. There were no signs of malignancy. The results of the study indicate that cASCs are similar to hASCs and are safe for therapeutic applications in a canine model. The proposed methodology for ASC preparation on a non-routine basis, which includes individually optimised cell culture conditions and offers risk-adapted treatment, could be used for future personalised off-the-shelf therapies, for example, in myocardial infarction or stroke.