Two-stage separation of immature and mature T lymphocytes from myeloerythroid clonogenic bone marrow cells by apheresis and elutriation centrifugation

Abstract

Because graft-versus-host disease remains a major complication in allogeneic bone marrow transplantation, a number of techniques capable of removing mature T lymphocytes from bone marrow cells have been attempted. The authors describe a simple two-step procedure using counterflow centrifugation elutriation (CCE) that eliminated 95 to 98 percent of the mature T lymphocytes and greater than 97 percent of the T lymphocyte colony-forming units (CFU-T) while concentrating the bone marrow myeloid colony-forming cells. Viability was greater than 98 percent, and 72 to 98 percent of the total cells separated were recovered. Lymphocyte depletion was substantiated by both morphologic and phenotypic criteria using monoclonal antibodies to T lymphocytes, as well as by responsiveness in mixed-lymphocyte cultures and to mitogens. In addition, this technique separated the lymphoid colony-forming cells from the larger myeloid colony-forming cells. It was concluded that this simple two-step CCE procedure can be used to separate T lymphocytes and CFU-T from myeloid colony-forming cells and offers a means of purging T lymphocytes from large numbers of marrow cells that may be required for human allogeneic bone marrow transplantation.