MegaCult™-C Complete Kit Without Cytokines

Complete kit for human CFU-Mk assays

MegaCult™-C Complete Kit Without Cytokines

Complete kit for human CFU-Mk assays

Catalog #
04970_C
Complete kit for human CFU-Mk assays
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What's Included

  • MegaCult™-C Complete Kit Without Cytokines (Catalog #04970)
    • Collagen Solution, 35 mL (Catalog #04902)
    • MegaCult™-C Medium Without Cytokines, 24 x 1.7 mL (Catalog #04900)
    • Double Chamber Slide Kit (Catalog #04963)
    • MegaCult™-C Staining Kit for CFU-Mk (Catalog #04962)

Overview

MegaCult™-C Complete Kit Without Cytokines includes all reagents (except cytokines) required for colony-forming unit (CFU) assays of megakaryocyte progenitor cells (CFU-Mk) in a collagen-based culture medium, including the detection of human CFU-Mk colonies by immunocytochemical staining. The addition of appropriate cytokines, as chosen by the user, is required.
Subtype
Semi-Solid Media, Specialized Media
Cell Type
Hematopoietic Stem and Progenitor Cells
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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Safety Data Sheet 1
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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 6
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Safety Data Sheet 7
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Safety Data Sheet 8
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Safety Data Sheet 9
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04970
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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

Frequently Asked Questions

Why is the MegaCult™-C formulation serum free?

MegaCult™-C is formulated without FBS to avoid inhibition of CFU-Mk growth by TGF beta and Platelet Factor-4, which are often present in the serum.

Why use semi-solid media?

Semi-solid media (such as methylcellulose-based or collagen-based) allow the clonal progeny of a single progenitor cell to stay together so you can recognize distinct colonies.

Publications (27)

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

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.
Application of human CFU-Mk assay to predict potential thrombocytotoxicity of drugs. Pessina A et al. Toxicology in vitro : an international journal published in association with BIBRA 2009 FEB

Abstract

Megakaryocytopoiesis gives rise to platelets by proliferation and differentiation of lineage-specific progenitors, identified in vitro as Colony Forming Unit-Megakaryocytes (CFU-Mk). The aim of this study was to refine and optimize the in vitro Standard Operating Procedure (SOP) of the CFU-Mk assay for detecting drug-induced thrombocytopenia and to prevalidate a model for predicting the acute exposure levels that cause maximum tolerated decreases in the platelets count, based on the correlation with the maximal plasma concentrations (C max) in vivo. The assay was linear under the SOP conditions, and the in vitro endpoints (percentage of colonies growing) were reproducible within and across laboratories. The protocol performance phase was carried out testing 10 drugs (selected on the base of their recognised or potential in vivo haematotoxicity, according to the literature). Results showed that a relationship can be established between the maximal concentration in plasma (C max) and the in vitro concentrations that inhibited the 10-50-90 percent of colonies growth (ICs). When C max is lower than IC10, it is possible to predict that the chemicals have no direct toxicity effect on CFU-Mk and could not induce thrombocytopenia due to bone marrow damage. When the C max is higher than IC90 and/or IC50, thrombocytopenia can occur due to direct toxicity of chemicals on CFU-Mk progenitors.
Incomplete restoration of Mpl expression in the mpl-/- mouse produces partial correction of the stem cell-repopulating defect and paradoxical thrombocytosis. Lannutti BJ et al. Blood 2009 FEB

Abstract

Expression of Mpl is restricted to hematopoietic cells in the megakaryocyte lineage and to undifferentiated progenitors, where it initiates critical cell survival and proliferation signals after stimulation by its ligand, thrombopoietin (TPO). As a result, a deficiency in Mpl function in patients with congenital amegakaryocytic thrombocytopenia (CAMT) and in mpl(-/-) mice produces profound thrombocytopenia and a severe stem cell-repopulating defect. Gene therapy has the potential to correct the hematopoietic defects of CAMT by ectopic gene expression that restores normal Mpl receptor activity. We rescued the mpl(-/-) mouse with a transgenic vector expressing mpl from the promoter elements of the 2-kb region of DNA just proximal to the natural gene start site. Transgene rescued mice exhibit thrombocytosis but only partial correction of the stem cell defect. Furthermore, they show very low-level expression of Mpl on platelets and megakaryocytes, and the transgene-rescued megakaryocytes exhibit diminished TPO-dependent kinase phosphorylation and reduced platelet production in bone marrow chimeras. Thrombocytosis is an unexpected consequence of reduced Mpl expression and activity. However, impaired TPO homeostasis in the transgene-rescued mice produces elevated plasma TPO levels, which serves as an unchecked stimulus to drive the observed excessive megakaryocytopoiesis.

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