Liu H et al. (DEC 2006)
Biomaterials 27 36 6004--14
Three-dimensional culture for expansion and differentiation of mouse embryonic stem cells.
Differentiation of embryonic stem (ES) cells typically requires cell-cell aggregation in the form of embryoid bodies (EBs). This process is not very well controlled and final cell numbers can be limited by EB agglomeration and the inability to drive differentiation towards a desired cell type. This study compares three-dimensional (3D) fibrin culture to conventional two-dimensional (2D) suspension culture and to culture in a semisolid methylcellulose medium solution. Two types of fibrin culture were evaluated,including a PEGylated fibrin gel. PEGylation with a difunctional PEG derivative retarded fibrinogen migration during through sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) as a result of crosslinking,similarly,degradation was slowed in the PEGylated gel. ES cell proliferation was higher in both the fibrin and PEGylated fibrin gels versus 2D and methylcellulose controls. FACS analysis and real-time-PCR revealed differences in patterns of differentiation for the various culture systems. Culture in PEGylated fibrin or methylcellulose culture demonstrated features characteristic of less extensive differentiation relative to fibrin and 2D culture as evidenced by the transcription factor Oct-4. Fibrin gels showed gene and protein expression similar to that in 2D culture. Both fibrin and 2D cultures demonstrated statistically greater cell numbers positive for the vascular mesoderm marker,VE-cadherin.
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J. E. Choi et al. (jun 2022)
STAR protocols 3 2 101389
Isolation of human and mouse myeloid-derived suppressor cells for metabolic analysis.
Metabolic reprogramming is associated with myeloid-derived suppressor cell (MDSC) immunosuppressive function. Here,we outline the process for acquiring MDSCs from human and murine sources for subsequent analysis of fatty acid oxidation,oxidative phosphorylation,and glycolysis using the Seahorse XFe 96 Analyzer. Murine MDSCs can be isolated directly from tumor-bearing mice or derived through IL-6 and GM-CSF culture of bone marrow cells from non-tumor-bearing mice. To generate human MDSCs,peripheral blood mononuclear cells (PBMCs) can be cultured with IL-6 and GM-CSF. For complete details on the use and execution of this protocol,please refer to Mohammadpour et al. (2021).
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Aldehyde dehydrogenase 1a1 is dispensable for stem cell function in the mouse hematopoietic and nervous systems.
High levels of aldehyde dehydrogenase (ALDH) activity have been proposed to be a common feature of stem cells. Adult hematopoietic,neural,and cancer stem cells have all been reported to have high ALDH activity,detected using Aldefluor,a fluorogenic substrate for ALDH. This activity has been attributed to Aldh1a1,an enzyme that is expressed at high levels in stem cells and that has been suggested to regulate stem cell function. Nonetheless,Aldh1a1 function in stem cells has never been tested genetically. We observed that Aldh1a1 was preferentially expressed in mouse hematopoietic stem cells (HSCs) and expression increased with age. Hematopoietic cells from Aldh1a1-deficient mice exhibited increased sensitivity to cyclophosphamide in a non-cell-autonomous manner,consistent with its role in cyclophosphamide metabolism in the liver. However,Aldh1a1 deficiency did not affect hematopoiesis,HSC function,or the capacity to reconstitute irradiated recipients in young or old adult mice. Aldh1a1 deficiency also did not affect Aldefluor staining of hematopoietic cells. Finally,Aldh1a1 deficiency did not affect the function of stem cells from the adult central or peripheral nervous systems. Aldh1a1 is not a critical regulator of adult stem cell function or Aldefluor staining in mice.
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Ibarra I et al. (DEC 2007)
Genes & development 21 24 3238--43
A role for microRNAs in maintenance of mouse mammary epithelial progenitor cells.
microRNA (miRNA) expression profiles are often characteristic of specific cell types. The mouse mammary epithelial cell line,Comma-Dbeta,contains a population of self-renewing progenitor cells that can reconstitute the mammary gland. We purified this population and determined its miRNA signature. Several microRNAs,including miR-205 and miR-22,are highly expressed in mammary progenitor cells,while others,including let-7 and miR-93,are depleted. Let-7 sensors can be used to prospectively enrich self-renewing populations,and enforced let-7 expression induces loss of self-renewing cells from mixed cultures.
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产品类型：
产品号#：
01700
01705
01702
产品名：
ALDEFLUOR™工具
ALDEFLUOR™ DEAB试剂
ALDEFLUOR™测定缓冲液
文献
Haddad EA et al. (SEP 2009)
Journal of immunology (Baltimore,Md. : 1950) 183 6 3608--15
An accessory role for B cells in the IL-12-induced activation of resting mouse NK cells.
IL-12 is a potent proinflammatory cytokine. The effects of IL-12 are thought to be mediated by IFN-gamma production by NK,NKT,and T cells. In this study,we show that although IL-12 stimulates NK and NK1.1(+) T cells in bulk mouse splenocytes,it does not significantly stimulate purified NK cells,indicating that other cells are required. IL-12 stimulates T cell-deficient spleen cells and those depleted of macrophages. Unexpectedly,the depletion of dendritic cells also has little effect on the stimulation of spleen cells with IL-12. In contrast,B cell depletion almost completely inhibits IL-12-induced IFN-gamma production and B cell-deficient spleen cells are poorly stimulated with IL-12. Furthermore,purified NK cells are stimulated with IL-12 in the presence of purified B cells. Thus,B cells are necessary and also sufficient for the stimulation of purified NK cells with IL-12. Whereas spleen cells from IL-18-deficient mice are not stimulated with IL-12,NK cells purified from IL-18-deficient mice are stimulated with IL-12 in the presence of wild-type (WT) B cells,and WT NK cells are not stimulated with IL-12 in the presence of IL-18-deficient B cells. Cell contact between B and NK cells is also required for IL-12-induced IFN-gamma production. Finally,B cell-deficient mice injected with IL-12 produce significantly less IFN-gamma and IL-18 in the sera than WT mice do. Thus,stimulation of NK cells with IL-12 requires B cell cooperation in vitro as well as in vivo.
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Aguila JR et al. (JUL 2011)
Blood 118 3 576--85
SALL4 is a robust stimulator for the expansion of hematopoietic stem cells.
HSCs are rare cells that have the unique ability to self-renew and differentiate into cells of all hematopoietic lineages. The lack of donors and current inability to rapidly and efficiently expand HSCs are roadblocks in the development of successful cell therapies. Thus,the challenge of ex vivo human HSC expansion remains a fertile and critically important area of investigation. Here,we show that either SALL4A- or SALL4B-transduced human HSCs obtained from the mobilized peripheral blood are capable of rapid and efficient expansion ex vivo by textgreater10 000-fold for both CD34(+)/CD38(-) and CD34(+)/CD38(+) cells in the presence of appropriate cytokines. We found that these cells retained hematopoietic precursor cell immunophenotypes and morphology as well as normal in vitro or vivo potential for differentiation. The SALL4-mediated expansion was associated with enhanced stem cell engraftment and long-term repopulation capacity in vivo. Also,we demonstrated that constitutive expression of SALL4 inhibited granulocytic differentiation and permitted expansion of undifferentiated cells in 32D myeloid progenitors. Furthermore,a TAT-SALL4B fusion rapidly expanded CD34(+) cells,and it is thus feasible to translate this study into the clinical setting. Our findings provide a new avenue for investigating mechanisms of stem cell self-renewal and achieving clinically significant expansion of human HSCs.
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EasySep™小鼠TIL（CD45）正选试剂盒
Cryopreservation Media for Stem Cell Research
文献
Positive Selection of Hematopoietic Stem and Progenitor Cells from Adult Mouse Bone Marrow
50:59
线上研讨会Gene Targeting in Hematopoietic Stem Cells for Basic and Translational Research发布日期: 11/28/18
技术窍门Genome Editing of Human CD34+ Hematopoietic Stem and Progenitor Cells Using the ArciTect™ CRISPR-Cas9 System and StemSpan™ Media
科学海报Optimized Media and Workflow for the Expansion of Human Pluripotent Stem Cells as Aggregates in Suspension Cultures
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