Mesenchymal stem cell (MSC) is the ideal cell source in regenerative medicine. The cell has multipotent and proliferation, and these features were useful for the therapeutic material in defective tissue. Separation and purification of MSC from living body is essential step for autologous cell plantation and tissue engineering. FACS or Ficoll methods generally separated the cells, but the isolated cells are not homogeneous. The therapeutic effects of target tissue using the cells were reported in clinical research. To establish the MSC plantation as a clinical treatment with safety, it is necessary to develop the isolation procedure for separating the homogeneous cells. In this research, we develop a cell separation column that was immobilized with antibody against MSC. The cell injected to the column roll on the surface under the flow condition like a rolling adhesion of the leukocyte in the blood vessel. Polyacrylic acid was grafted on the surface of the polyethylene or silicone tube by ozone-induce graft polymerization. The anti-human CD34 antibody was immobilized on the surface. The cell suspension passed through the column, and the number and surface marker pattern of the cells in each elution fraction were analyzed. Mouse bone marrow (BM) was isolated from mice by flushing the marrow cavities. The MSC's were prepared by the BM cultured on plastic dish. The cell suspension passed through the antibody-immobilized column, 10% of the whole cells were observed as delayed fraction. The fraction was found to consist of the cells with high density of surface marker. In contrast, the cell suspension passed the control column, the delayed fraction was not observed. These results suggested that the cells in delayed fraction rolled on the column surface in the marker specific manner under the flow condition, and that system are useful to isolate the cells based on the density of surface markers.