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Erythroid Progenitor Reprogramming Kit

Integrated workflow containing tools to isolate and expand erythroid progenitor cells from peripheral blood and their subsequent reprogramming to iPS cells
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产品优势


  • Optimized for enrichment, expansion and reprogramming of erythroid progenitor cells expanded from peripheral blood samples

  • Improved reprogramming efficiency and higher frequency of iPS cell colonies, compared to traditional hES cell medium

  • Rapid emergence of large colonies with high quality iPS cell-like morphology facilitates identification and subcloning

  • Seamlessly integrates with TeSR™ and STEMdiff™ products for downstream maintenance and differentiation of iPS cell lines

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    概述
    实验数据
    产品说明书及文档
    应用领域
    相关材料与文献
    The Erythroid Progenitor Reprogramming Kit contains an integrated set of tools and reagents to enrich, expand and reprogram erythroid progenitor cells from peripheral blood.

    Data Figures

    Depletion of T-Cells and B-Cells From Whole Blood With RosetteSep™ Human Progenitor Cell Basic Pre-Enrichment Cocktail and SepMate™ Tubes

    Figure 1. Depletion of T-Cells and B-Cells from Whole Blood with RosetteSep™ Human Progenitor Cell Basic Pre-Enrichment Cocktail and SepMate™ Tubes

    (A) In the donor sample shown above, T-cells (CD3+) represent approximately 21.2% of the PBMC fraction while B-cells (CD19+) are present at 3.4%. (B) The addition of the RosetteSep™ Human Progenitor Cell Basic Pre-Enrichment cocktail efficiently depletes the T- and B-cell population to <1% of the enriched cell fraction.

    Expansion of Erythroid Progenitor Cells Isolated From Peripheral Blood Using StemSpan™ SFEM II and StemSpan™ Erythroid Expansion Supplement

    Figure 2. Expansion of Erythroid Progenitor Cells Isolated from Peripheral Blood Using StemSpan™ SFEM II and StemSpan™ Erythroid Expansion Supplement

    TNC: total nucleated cell, Average shown in bold (range).

    Erythroid Progenitor Cells are Expanded in StemSpan™ SFEM II With Erythroid Expansion Supplement

    Figure 3. Erythroid Progenitor Cells Are Expanded in StemSpan™ SFEM II with Erythroid Expansion Supplement

    (A) Isolated PBMCs were expanded for seven days and then examined by flow cytometry for erythroid progenitor cells, T-cells and B-cells. Representative plots illustrate that erythroid progenitor cells (GlyA+CD71+) are enriched after seven days, though some T-cells (CD3+) and B-cells (CD19+) remain. (B) Use of the RosetteSep™ cocktail to deplete lineage-committed cells leads to increased purity of the expanded erythroid progenitors and little/no contaminating lymphoid cells. Note: same donor sample used for A and B.

    Schematic of ReproTeSR™ Reprogramming Timeline

    Figure 4. Schematic of ReproTeSR™ Reprogramming Timeline

    ReproTeSR™ is used during the entire induction phase of reprogramming (day 3 to 21). On days 3 and 5, ReproTeSR™ is added to StemSpan™ growth media (in a fed-batch manner) to facilitate attachment of transfected cells. Attached cells are further cultured in ReproTeSR™ with daily full media changes until putative iPS cell colonies emerge (days 21-28). iPS cell colonies can then be isolated and propagated in TeSR™ media. (mTeSR™1, TeSR™2, TeSR™-E8™).

    Blood Cell Reprogramming Efficiencies Are Higher in ReproTeSR™ Medium Compared to in hESC Medium

    Figure 5. Blood Cell Reprogramming Efficiencies Are Higher in ReproTeSR™ Medium Compared to in hESC Medium

    Efficiency of reprogramming (A) erythroid cells, or (B) CD34+ cells using episomal reprogramming vectors is higher in ReproTeSR™ medium compared to in KOSR-containing hESC medium. Data shown are mean +/- SEM, erythroid cells n=4, CD34+ cells n=5.

    Generation of iPS Cells From 1mL of Peripheral Blood

    Figure 6. Generation of iPS Cells from 1mL of Peripheral Blood

    Starting from 1mL of PB, PBMCs were enriched, erythroid progenitors were expanded and reprogrammed in ReproTeSR™. (A) Approximately 75 iPS-like colonies that were positive for alkaline phosphatase expression (blue) were generated. (B-C) iPS cell colonies exhibit compact ES-like morphology with defined borders and high nuclear to cytoplasmic ratio. Representative images of generated iPS cell colonies taken at 20X (B) and 400X (C) magnification are shown.

    Reprogramming Efficiency of CD34+ and Erythroid Progenitor Cells With ReproTeSR™

    Figure 7. Reprogramming Efficiency of CD34+ and Erythroid Progenitor Cells With ReproTeSR™

    Average values in bold (range).

    Protocols and Documentation

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

    Document Type
    Product Name
    Catalog #
    Lot #
    Language
    Document Type
    Technical Manual
    Product Name
    Erythroid Progenitor Reprogramming Kit
    Catalog #
    05924
    Lot #
    All
    Language
    English
    Document Type
    Safety Data Sheet 1
    Product Name
    Erythroid Progenitor Reprogramming Kit
    Catalog #
    05924
    Lot #
    All
    Language
    English
    Document Type
    Safety Data Sheet 2
    Product Name
    Erythroid Progenitor Reprogramming Kit
    Catalog #
    05924
    Lot #
    All
    Language
    English
    Document Type
    Safety Data Sheet 3
    Product Name
    Erythroid Progenitor Reprogramming Kit
    Catalog #
    05924
    Lot #
    All
    Language
    English
    Document Type
    Safety Data Sheet 4
    Product Name
    Erythroid Progenitor Reprogramming Kit
    Catalog #
    05924
    Lot #
    All
    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.

    Research Area
    Workflow Stages

    Resources and Publications

    Educational Materials (4)

    An Integrated Workflow for Reprogramming Blood Cells
    Brochure
    An Integrated Workflow for Reprogramming Blood Cells
    hPSC Differentiation: Tools for Pluripotent Stem Cell-Derived Research
    Brochure
    hPSC Differentiation: Tools for Pluripotent Stem Cell-Derived Research
    Products for Human Pluripotent Stem Cells
    Brochure
    Products for Human Pluripotent Stem Cells
    Defined and Xeno-Free Medium for Reprogramming Blood-Derived CD34+ Cells or Erythroid Cells
    Scientific Poster
    Defined and Xeno-Free Medium for Reprogramming Blood-Derived CD34+ Cells or Erythroid Cells

    Publications (1)

    Development of induced pluripotent stem cells from a patient with hypertrophic cardiomyopathy who carries the pathogenic myosin heavy chain 7 mutation p.Arg403Gln. M. Holliday et al. Stem cell research 2018

    Abstract

    Hypertrophic cardiomyopathy (HCM) is an inherited cardiomyopathy characterized by left ventricular hypertrophy ≥15 mm in the absence of loading conditions. HCM has a prevalence of up to one in 200, and can result in significant adverse outcomes including heart failure and sudden cardiac death. An induced pluripotent stem cell (iPSC) line was generated from peripheral blood mononuclear cells obtained from the whole blood of a 38-year-old female patient with HCM in which genetic testing identified the well-known pathogenic p.Arg403Gln mutation in myosin heavy chain 7. iPSCs express pluripotency markers, demonstrate trilineage differentiation capacity, and display a normal 46,XX female karyotype. This resource will allow further assessment of the pathophysiological development of HCM.
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