Purification of human peripheral blood colony forming cells (CFUC).
A 10-fold enrichment of colony forming cells (CFUC) from single donor platelet-apheresis residues and from 70-120 ml of peripheral blood of normal donors was achieved by sequential sedimentation on Ficoll-diatrizoate, depletion of cells adherent to plastic, and depletion of cells rosetting with sheep red blood cells (T lymphocytes). Culture of 5 x 10(5) unfractionated mononuclear cells yielded 9 +/- 3 colonies and mononuclear cells depleted of adherent cells and T lymphocytes yielded 53 +/- 6 colonies. The mononuclear cell fraction depleted of adherent cells and T lymphocytes was further enriched for CFUC by isopycnic sedimentation of Percoll gradients. Cells recovered in the 1.0063-1.065 g/cm3 density layer of the gradient formed 146 +/- 9 colonies in culture. The mononuclear cells depleted of adherent cells and T lymphocytes were also enriched for CFUC by depletion of Fc-receptor positive cells using an immune sheep red blood cell rosette sedimentation technique. Cultures of the Fc-receptor depleted fractions yielded 107 +/- 12 colonies, while the Fc-receptor enriched fraction yielded only 2 +/- 1 colonies. CFUC appear to lack surface membrane receptors for sheep erythrocytes and the Fc portion of immunoglobulin as well as the ability to adhere to plastic.
D002448:Cell Adhesion; D002469:Cell Separation / Q000379:methods*; D002478:Cells, Cultured; D002500:Centrifugation, Isopycnic; D003470:Culture Media; D006412:Hematopoietic Stem Cells / Q000276:immunology*; D006801:Humans; D007141:Immunoglobulin Fc Fragments; D007158:Immunologic Techniques; D008212:Lymphocyte Depletion; D010983:Plateletpheresis; D013601:T-Lymphocytes / Q000276:immunology
D003470:Culture Media; D007141:Immunoglobulin Fc Fragments