Functional Study and Detection of HLA‐ D Products on Fractionated Human Bone Marrow Cells

Abstract

Bone marrow cells from nine normal human volunteers, obtained from the Iliac crest, were used in this work for antigen determination and functional studies. The bone marrow aspirated cells were sequentially separated: elimination of erythrocytes, granulocytes and monocytes was achieved by Ficoll-Isopaque centrifugation followed by plastic adherence. Purified bone marrow cells were finally separated by size using velocity sedimentation. The slow sedimenting small cells were shown to be mainly T lymphocytes, probably of blood origin. The medium sized bone marrow cells were shown to contain myeloid precursors (CFu-c). Large immature cells were in cycle actively synthesizing DNA molecules.HLA-D and HLA-DR detections on the fractionated cells were performed using three technique: fluorescence with specific anti HLA-DR allo and xeno antisera; primed lymphocyte typing (PLT) with anti HLA-DR monospecific in vitro primed lymphocytes and detection of the HLA-D stimulating product using the bone marrow fractionated cells as stimulators in a mixed leukocyte culture. Concordant results were obtained with the three techniques. Lymphocytes in the bone marrow express HLA-D. products as peripheral lymphocytes. Bone marrow fractions depleted of lymphocytes and monocytes also contain approximately 20% of cells expressing HLA-D products. The meaning of the expression of HLA-D products on immature precursors non-lymphoid cells is discussed.