MegaCult™-C Complete Kit with Cytokines

Complete kit for human CFU-Mk assays
MegaCult™-C Complete Kit with Cytokines

Complete kit including collagen, media, staining kit and chamber slides for human CFU-Mk assays

1 Kit
Catalog # 04971
5,713 USD
Collagen Solution

For preparation of collagen gels and for coating cell culture surfaces

35 mL
Catalog # 04902
519 USD
MegaCult™-C Medium with Cytokines

Medium with cytokines

24 x 2 mL
Catalog # 04901
1,386 USD
Double Chamber Slide Kit

For the culture, fixation and staining of collagen-based cultures

1 Kit
Catalog # 04963
1,517 USD
MegaCult™-C Staining Kit for CFU-Mk

Staining kit for human CFU-Mk assays

1 Kit
Catalog # 04962
2,520 USD

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.
Components
  • 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)
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

Scientific Resources

Product Documentation

Document Type Product Name Catalog # Lot # Language
Document Type
Product Information Sheet
Product Name
MegaCult™-C Complete Kit with Cytokines
Catalog #
04971
Lot #
All
Language
English
Document Type
Product Information Sheet
Product Name
Collagen Solution
Catalog #
04902
Lot #
All
Language
English
Document Type
Product Information Sheet
Product Name
MegaCult™-C Medium with Cytokines
Catalog #
04901
Lot #
All
Language
English
Document Type
Product Information Sheet
Product Name
Double Chamber Slide Kit
Catalog #
04963
Lot #
All
Language
English
Document Type
Product Information Sheet
Product Name
MegaCult™-C Staining Kit for CFU-Mk
Catalog #
04962
Lot #
All
Language
English
Document Type
Manual
Product Name
MegaCult™-C Complete Kit with Cytokines
Catalog #
04971
Lot #
All
Language
English
Document Type
Safety Data Sheet 1
Product Name
MegaCult™-C Complete Kit with Cytokines
Catalog #
04971
Lot #
All
Language
English
Document Type
Safety Data Sheet 2
Product Name
MegaCult™-C Complete Kit with Cytokines
Catalog #
04971
Lot #
All
Language
English
Document Type
Safety Data Sheet 3
Product Name
MegaCult™-C Complete Kit with Cytokines
Catalog #
04971
Lot #
All
Language
English
Document Type
Safety Data Sheet 4
Product Name
MegaCult™-C Complete Kit with Cytokines
Catalog #
04971
Lot #
All
Language
English
Document Type
Safety Data Sheet 5
Product Name
MegaCult™-C Complete Kit with Cytokines
Catalog #
04971
Lot #
All
Language
English
Document Type
Safety Data Sheet 6
Product Name
MegaCult™-C Complete Kit with Cytokines
Catalog #
04971
Lot #
All
Language
English
Document Type
Safety Data Sheet 7
Product Name
MegaCult™-C Complete Kit with Cytokines
Catalog #
04971
Lot #
All
Language
English
Document Type
Safety Data Sheet 8
Product Name
MegaCult™-C Complete Kit with Cytokines
Catalog #
04971
Lot #
All
Language
English
Document Type
Safety Data Sheet 9
Product Name
MegaCult™-C Complete Kit with Cytokines
Catalog #
04971
Lot #
All
Language
English
Document Type
Safety Data Sheet
Product Name
Collagen Solution
Catalog #
04902
Lot #
All
Language
English
Document Type
Safety Data Sheet
Product Name
MegaCult™-C Medium with Cytokines
Catalog #
04901
Lot #
All
Language
English
Document Type
Safety Data Sheet
Product Name
MegaCult™-C Staining Kit for CFU-Mk
Catalog #
04962
Lot #
All
Language
English

Product 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.

Data and Publications

Publications (31)

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

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.
Blood 2010 SEP Serum response factor is an essential transcription factor in megakaryocytic maturation. Halene S et al.

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.
Blood 2009 SEP c-Myc is a target of RNA-binding motif protein 15 in the regulation of adult hematopoietic stem cell and megakaryocyte development. Niu C et al.

Abstract

RNA-binding motif protein 15 (RBM15) is involved in the RBM15-megakaryoblastic leukemia 1 fusion in acute megakaryoblastic leukemia. Although Rbm15 has been reported to be required for B-cell differentiation and to inhibit myeloid and megakaryocytic expansion, it is not clear what the normal functions of Rbm15 are in the regulation of hematopoietic stem cell (HSC) and megakaryocyte development. In this study, we report that Rbm15 may function in part through regulation of expression of the proto-oncogene c-Myc. Similar to c-Myc knockout (c-Myc-KO) mice, long-term (LT) HSCs are significantly increased in Rbm15-KO mice due to an apparent LT-HSC to short-term HSC differentiation defect associated with abnormal HSC-niche interactions caused by increased N-cadherin and beta(1) integrin expression on mutant HSCs. Both serial transplantation and competitive reconstitution capabilities of Rbm15-KO LT-HSCs are greatly compromised. Rbm15-KO and c-Myc-KO mice also share related abnormalities in megakaryocyte development, with mutant progenitors producing increased, abnormally small low-ploidy megakaryocytes. Consistent with a possible functional interplay between Rbm15 and c-Myc, the megakaryocyte increase in Rbm15-KO mice could be partially reversed by ectopic c-Myc. Thus, Rbm15 appears to be required for normal HSC-niche interactions, for the ability of HSCs to contribute normally to adult hematopoiesis, and for normal megakaryocyte development; these effects of Rbm15 on hematopoiesis may be mediated at least in part by c-Myc.
Blood 2009 SEP miR-34a contributes to megakaryocytic differentiation of K562 cells independently of p53. Navarro F et al.

Abstract

The role of miRNAs in regulating megakaryocyte differentiation was examined using bipotent K562 human leukemia cells. miR-34a is strongly up-regulated during phorbol ester-induced megakaryocyte differentiation, but not during hemin-induced erythrocyte differentiation. Enforced expression of miR-34a in K562 cells inhibits cell proliferation, induces cell-cycle arrest in G(1) phase, and promotes megakaryocyte differentiation as measured by CD41 induction. miR-34a expression is also up-regulated during thrombopoietin-induced differentiation of CD34(+) hematopoietic precursors, and its enforced expression in these cells significantly increases the number of megakaryocyte colonies. miR-34a directly regulates expression of MYB, facilitating megakaryocyte differentiation, and of CDK4 and CDK6, to inhibit the G(1)/S transition. However, these miR-34a target genes are down-regulated rapidly after inducing megakaryocyte differentiation before miR-34a is induced. This suggests that miR-34a is not responsible for the initial down-regulation but may contribute to maintaining their suppression later on. Previous studies have implicated miR-34a as a tumor suppressor gene whose transcription is activated by p53. However, in p53-null K562 cells, phorbol esters induce miR-34a expression independently of p53 by activating an alternative phorbol ester-responsive promoter to produce a longer pri-miR-34a transcript.
Blood 2009 SEP Platelet factor 4 regulates megakaryopoiesis through low-density lipoprotein receptor-related protein 1 (LRP1) on megakaryocytes. Lambert MP et al.

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.
Blood 2009 MAR Role for MKL1 in megakaryocytic maturation. Cheng E-C et al.

Abstract

Megakaryoblastic leukemia 1 (MKL1), identified as part of the t(1;22) translocation specific to acute megakaryoblastic leukemia, is highly expressed in differentiated muscle cells and promotes muscle differentiation by activating serum response factor (SRF). Here we show that Mkl1 expression is up-regulated during murine megakaryocytic differentiation and that enforced overexpression of MKL1 enhances megakaryocytic differentiation. When the human erythroleukemia (HEL) cell line is induced to differentiate with 12-O-tetradecanoylphorbol 13-acetate, overexpression of MKL1 results in an increased number of megakaryocytes with a concurrent increase in ploidy. MKL1 overexpression also promotes megakaryocytic differentiation of primary human CD34(+) cells cultured in the presence of thrombopoietin. The effect of MKL1 is abrogated when SRF is knocked down, suggesting that MKL1 acts through SRF. Consistent with these findings in human cells, knockout of Mkl1 in mice leads to reduced platelet counts in peripheral blood, and reduced ploidy in bone marrow megakaryocytes. In conclusion, MKL1 promotes physiologic maturation of human and murine megakaryocytes.
View All Publications

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