造血干细胞
The definitive hematopoietic system is made up of all adult blood cell types including megakaryocytes, erythrocytes and cells of the myeloid and lymphoid lineages. All of these cells are derived from multipotent hematopoietic stem cells (HSCs) through a succession of precursors with progressively limited potential. Hematopoietic stem cells (HSCs) are tissue-specific stem cells that exhibit remarkable self-renewal capacity and are responsible for the life-long maintenance of hematopoietic system. HSCs are rare cells that reside in adult bone marrow where hematopoiesis is continuously taking place, and they can also be found in cord blood, fetal liver, adult spleen and peripheral blood.
R&D Systems offers several products for assessing hematopoietic lineage cells including serum-free media, lineage depletion antibodies and kits, and reagents for performing colony forming cell (CFC) assays.
Serum-Free Media for Hematopoietic Cell Culture
The StemXVivo Serum-Free Dendritic Cell Base Media and T Cell Base Media are formulated and optimized for the culture of human dendritic cells and T cells, respectively. These media must be supplemented with cytokines/growth factors for the desired cell culture application. The cytokine/growth factor combinations used depends upon the experimental design of each researcher.
For a protocol highlighting the use of the StemXVivo Human Dendritic Cell Base Media, please visit our website at www.RnDSystems.com/go/DCProtocol. For a protocol highlighting the use of StemXVivo Serum-free T Cell Base Media, please visit our website at www.RnDSystems.com/go/ExVivoTCellProtocol
Dendritic Cell Culture and Maturation from CD14+ Monocytes. A: LPS-matured monocyte-derived dendritic cells were obtained after a 9-day culture of CD14-enriched monocytes in StemXVivo Serum-Free Dendritic Cell Base Media (Catalog # CCM003) supplemented with GM-CSF (Catalog # 215-GM) and IL-4 (Catalog # 204-IL). B: Dendritic cells cultured for 7 days were stained with the mature dendritic cell marker CD83 before (gray line) and after a two day treatment with LPS (black line). Isotype-matched control staining is also shown (filled histogram).
Supplements for Hematopoietic Stem Cell Expansion
R&D Systems offers the Mouse Hematopoietic Cytokine Panel containing supplements that support the expansion of hematopoietic stem cells. This panel provides an economical alternative to purchasing each protein separately.
Hematopoietic Lineage Depletion
Mouse Hematopoietic Lineage Depletion Antibodies
Anti-mouse monoclonal antibodies can be used to efficiently bind lineage-committed bone marrow-derived cells. Antibodies included in the lineage depletion panel are optimized and contain sufficient amounts to process 1 x 109 bone marrow-derived cells. They can conveniently be used in conjunction with magnetic particle separation systems or flow cytometric cell sorting for the enrichment of uncommitted mesenchymal or hematopoietic stem cells.
Mouse Hematopoietic Cell Lineage Depletion Kit
The Mouse Hematopoietic Cell Lineage Depletion Kit is designed to be used in conjunction with R&D Systems MagCellect? cell enrichment system. Lineage committed cells (lineage positive) targeted for depletion include T cells, B cells, NK cells, monocytes/macrophages, granulocytes, and erythrocytes. The kit comes with reagents to process 1 x 109 cells. A protocol for using the Mouse Hematopoietic Lineage Depletion Kit can be found on pages 26-27.
Hematopoietic Lineage Depletion. Lineage marker reactivity on BALB/c bone marrow (BM) cells processed with the MagCellect Mouse Hematopoietic Cell Lineage Depletion Kit (Catalog # MAGM209). Histograms show reactivity of BM cells labeled with the cocktail of biotinylated antibodies included in the kit both before (green histogram) and after (open histogram) magnetic depletion. Lineage marker reactivity was detected using Streptavidin-PE.
Enrichment of CD117+ Cells. CD117 (c-kit) staining of bone marrow lineage negative cells isolated from BALB/c mice using the MagCellect Mouse Hematopoietic Cell Lineage Depletion Kit (Catalog # MAGM209). Histograms reflect reactivity of all viable cells with anti-CD117-PE (Catalog # FAB1356P; light green) or a matched isotype control antibody (open histogram). Inset shows CD117 staining of cells before depletion.
Methylcellulose-based Reagents for Colony Forming Cell Assays
The colony forming cell (CFC) assay, also known as the clonal culture assay, is the standard in vitro assay for quantitation of clonogenic progenitors present in human, murine, and primate bone marrow, umbilical cord blood, peripheral blood, and G-CSF mobilized peripheral blood. This assay relies on the ability of hematopoietic progenitors to proliferate and differentiate into distinct colonies in a semi-solid media in response to growth factor stimulation. The unique morphology of the resulting colonies enables enumeration and characterization of the progenitors. Since methylcellulose has replaced agar as the immobilizing agent used in the CFC assay, this assay is also known as the methylcellulose assay. Hematopoietic colony-forming assays may be used to evaluate potential toxic effects of new compounds and to determine maximum tolerated doses (MTD) and inhibitory concentration values (IC50).
Please see pages 28-31 for a protocol describing the use of methylcellulose-based reagents for colony forming cell assays.
*HSC001 and HSC002 do not contain any cytokines and will not support colony growth unless conditioned media, cytokines, or other culture supplements are added.
Recent references using Methylcellulose-based products from R&D Systems
Additional Selected Stem Cell Culture-related Products
Please see our website at www. RnDSystems.com/go/StemCell for a complete product listing.
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