FACS

Recent transplantation experiments have shown, that all blood cell lineages in the zebrafish develop from hematopoietic stem cells in the kidney. Analysis of these lineages by flow cytometry revealed surprisingly, that based on light scatter characteristics alone, four populations of hematopoietic lineages could be clearly separated (see Fig.1): erythroid cells (top left), myeloid cells (top right), precursor cells (bottom right) and lymphocytes (bottom left). These results were confirmed by cytospin and microscopic examination of the respective populations (see Fig.1 inserts). This technology allows for examination of steady state hematopoiesis in blood mutants. Furthermore, distinct populations can be sorted for transplantation purposes. The transplantation experiments can be refined by selecting subpopulations of cells within a light scatter gate by using lineage-specific GFP expression. In this way we were able to sort out for example immature B cells from the lymphoid population of the Rag-2-GFP transgenic kidney (Fig.2a,b), and mature T cells from the lymphoid population of the lck-GFP transgenic kidney (Fig.2c,d). Immature T cells were isolated from the lymphoid gate in the thymus using the Rag-2-promoter-GFP transgenic line (Fig. 2e,f), or all T cells by using the lck-promoter-GFP line (see Fig.2g,h).