Clonal T-cell colony formation in agar culture: an attractive assay to test the T-cell depletion from bone marrow.
Current studies suggest that the depletion of T-lymphocytes from donor marrow is an effective method for preventing acute graft-versus-host disease (GVHD) after allogeneic bone marrow transplantation in man. To deplete the T-lymphocytes from bone marrow cells we use either monoclonal anti-T-cell antibodies and complement or T101 ricin A-chain immunotoxin. Residual T-lymphocytes are analyzed by their capacity to form clonal T-cell colonies in the presence of phytohemagglutinin (PHA), accessory cells, and recombinant interleukin 2. The method is compared to immediate indirect immunofluorescence (iF) and thymidine incorporation by marrow cells stimulated by PHA. IF is not suitable for evaluating the depletion by immunotoxin, and the interpretation of thymidine incorporation is generally questionable. The results of the colony formation show that the sensitivity of the colony assay is close to that of iF when T cells are depleted by complement lysis, and the sensitivity of the colony assay is not dependent upon the depletion procedure. Therefore, the T-cell colony assay is a simple functional control for the quality of bone marrow T-cell depletion, especially for T-cell depletion by immunotoxin.
D000362:Agar; D001854:Bone Marrow Cells; D016026:Bone Marrow Transplantation; D003114:Colony-Forming Units Assay; D005455:Fluorescent Antibody Technique; D006086:Graft vs Host Disease / Q000517:prevention & control; D006801:Humans; D007958:Leukocyte Count; D013601:T-Lymphocytes / Q000166:cytology*; D013936:Thymidine / Q000378:metabolism
D000362:Agar; D013936:Thymidine