MegaCult™-C Complete Kit with Cytokines

Complete kit for human CFU-Mk assays

MegaCult™-C Complete Kit with Cytokines

Complete kit for human CFU-Mk assays

Catalog #
04971_C
Complete kit for human CFU-Mk assays
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What's Included

  • MegaCult™-C Complete Kit with Cytokines (Catalog #04971)
    • Collagen Solution, 35 mL (Catalog #04902)
    • MegaCult™-C Medium with Cytokines, 24 x 2 mL (Catalog #04901)
    • Double Chamber Slide Kit (Catalog #04963)
    • MegaCult™-C Staining Kit for CFU-Mk (Catalog #04962)

Overview

MegaCult™-C Complete Kit with Cytokines includes all reagents required for the optimal growth of megakaryocyte progenitors (CFU-Mk) within double chamber slides and for the detection of CFU-Mk colonies by immunocytochemical staining.

MegaCult™-C medium is optimized for the growth of CFU-Mk in human bone marrow, mobilized peripheral blood and cord blood samples. It is suitable for use with CD34+-enriched cells, mononuclear cells and cells isolated by other purification methods.
Contains
• Collagen Solution, 35 mL (Catalog #04902)
• MegaCult™-C Medium with Cytokines, 24 x 2 mL (Catalog #04901)
• Double Chamber Slide Kit, 1 Kit (Catalog #04963)
• MegaCult™-C Staining Kit for CFU-Mk, 1 Kit (Catalog #04962)
Subtype
Semi-Solid Media, Specialized Media
Cell Type
Hematopoietic Stem and Progenitor Cells
Species
Human
Application
Cell Culture, Colony Assay, Functional Assay
Brand
MegaCult
Area of Interest
Stem Cell Biology
Formulation
Serum-Free

Protocols and Documentation

Find supporting information and directions for use in the Product Information Sheet or explore additional protocols below.

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English
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Technical Manual
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English
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Safety Data Sheet 1
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English
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Safety Data Sheet 2
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Safety Data Sheet 3
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Safety Data Sheet 4
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Safety Data Sheet 5
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Safety Data Sheet 6
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Safety Data Sheet 7
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Safety Data Sheet 8
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English
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Safety Data Sheet 9
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English
Catalog #
04971
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Language
English

Applications

This product is designed for use in the following research area(s) as part of the highlighted workflow stage(s). Explore these workflows to learn more about the other products we offer to support each research area.

Resources and Publications

Publications (31)

Identification of unipotent megakaryocyte progenitors in human hematopoiesis. Miyawaki K et al. Blood 2017 MAR

Abstract

The developmental pathway for human megakaryocytes remains unclear and the definition of pure unipotent megakaryocyte progenitor is still controversial. Using single-cell transcriptome analysis, we have identified a cluster of cells within immature hematopoietic stem and progenitor cell populations that specifically express genes related to the megakaryocyte lineage. We used CD41 as a positive marker to identify these cells within the CD34(+)CD38(+)IL-3Rα(dim)CD45RA(-) common myeloid progenitor (CMP) population. These cells lacked erythroid and granulocyte/macrophage potential, but exhibited robust differentiation into the megakaryocyte lineage at a high frequency, both in vivo and in vitro The efficiency and expansion potential of these cells exceeded those of conventional bipotent megakaryocyte/erythrocyte progenitors. Accordingly, the CD41(+) CMP was defined as a unipotent megakaryocyte progenitor (MegP) that is likely to represent the major pathway for human megakaryopoiesis, independent of canonical megakaryocyte-erythroid lineage bifurcation. In the bone marrow of patients with essential thrombocythemia, the MegP population was significantly expanded in the context of a high burden of Janus kinase 2 mutations. Thus, the prospectively isolatable and functionally homogeneous human MegP will be useful for the elucidation of the mechanisms underlying normal and malignant human hematopoiesis.
Serum response factor is an essential transcription factor in megakaryocytic maturation. Halene S et al. Blood 2010 SEP

Abstract

Serum response factor (Srf) is a MADS-box transcription factor that is critical for muscle differentiation. Its function in hematopoiesis has not yet been revealed. Mkl1, a cofactor of Srf, is part of the t(1;22) translocation in acute megakaryoblastic leukemia, and plays a critical role in megakaryopoiesis. To test the role of Srf in megakaryocyte development, we crossed Pf4-Cre mice, which express Cre recombinase in cells committed to the megakaryocytic lineage, to Srf(F/F) mice in which functional Srf is no longer expressed after Cre-mediated excision. Pf4-Cre/Srf(F/F) knockout (KO) mice are born with normal Mendelian frequency, but have significant macrothrombocytopenia with approximately 50% reduction in platelet count. In contrast, the BM has increased number and percentage of CD41(+) megakaryocytes (WT: 0.41% ± 0.06%; KO: 1.92% ± 0.12%) with significantly reduced ploidy. KO mice show significantly increased megakaryocyte progenitors in the BM by FACS analysis and CFU-Mk. Megakaryocytes lacking Srf have abnormal stress fiber and demarcation membrane formation, and platelets lacking Srf have abnormal actin distribution. In vitro and in vivo assays reveal platelet function defects in KO mice. Critical actin cytoskeletal genes are down-regulated in KO megakaryocytes. Thus, Srf is required for normal megakaryocyte maturation and platelet production partly because of regulation of cytoskeletal genes.
Platelet factor 4 regulates megakaryopoiesis through low-density lipoprotein receptor-related protein 1 (LRP1) on megakaryocytes. Lambert MP et al. Blood 2009 SEP

Abstract

Platelet factor 4 (PF4) is a negative regulator of megakaryopoiesis, but its mechanism of action had not been addressed. Low-density lipoprotein (LDL) receptor-related protein-1 (LRP1) has been shown to mediate endothelial cell responses to PF4 and so we tested this receptor's importance in PF4's role in megakaryopoiesis. We found that LRP1 is absent from megakaryocyte-erythrocyte progenitor cells, is maximally present on large, polyploidy megakaryocytes, and near absent on platelets. Blocking LRP1 with either receptor-associated protein (RAP), an antagonist of LDL family member receptors, or specific anti-LRP1 antibodies reversed the inhibition of megakaryocyte colony growth by PF4. In addition, using shRNA to reduce LRP1 expression was able to restore megakaryocyte colony formation in bone marrow isolated from human PF4-overexpressing mice (hPF4(High)). Further, shRNA knockdown of LRP1 expression was able to limit the effects of PF4 on megakaryopoiesis. Finally, infusion of RAP into hPF4(High) mice was able to increase baseline platelet counts without affecting other lineages, suggesting that this mechanism is important in vivo. These studies extend our understanding of PF4's negative paracrine effect in megakaryopoiesis and its potential clinical implications as well as provide insights into the biology of LRP1, which is transiently expressed during megakaryopoiesis.

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