EXPERIMENTAL FIGURE 21-18 Functional analysis of hematopoietic stem cells by bone marrow transplantation. The two strains of mice used in this analysis are genetically identical except for the gene encoding a protein, termed Ly5, found on the surfaces of all nucleated blood cells, including all T and B lymphocytes, granulocytes, and monocytes. The proteins encoded by the two alleles of the gene, Ly5.1 and Ly5.2, can be detected by specific monoclonal antibodies. A recipient Ly5.2 mouse is lethally irradiated to kill all HSCs, then injected with stem cells purified from a Ly5.1 strain. Because the stem cells take weeks or months to produce differentiated blood cells, the recipient mouse will die unless it receives bone marrow progenitor cells from a genetically identical mouse (termed “supportive” cells) that will produce mature blood cells for the first few weeks after the transplant. At intervals after the transplant, blood or bone marrow is recovered and reacted with a blue-fluorescing monoclonal antibody to Ly5.1 and a red-fluorescing monoclonal antibody to Ly5.2. Mature blood cells that are descended from the donor stem cell are detected by FACS analysis, seen here as cells that fluoresce blue and not red. These cells can be sorted and stained with fluorescent antibodies specific for marker proteins found on different types of mature blood cells to show that a stem cell is indeed pluripotent, in that it can generate all types of lymphoid and myeloid cells.

[Courtesy Dr. Chengcheng (Alec) Zhang.]