• 我要登录|
  • 免费注册
    |
  • 我的丁香通
    • 企业机构:
    • 成为企业机构
    • 个人用户:
    • 个人中心
  • 移动端
    移动端
丁香通 logo丁香实验_LOGO
搜实验

    大家都在搜

      大家都在搜

        0 人通过求购买到了急需的产品
        免费发布求购
        发布求购
        点赞
        收藏
        wx-share
        分享

        Method for Purification of Human Hematopoietic Stem Cells by Flow Cytometry

        互联网

        1122
        Human hematopoietic stem cells (HSCs) and progenitors can be isolated by enriching for a rare cell population with a combination of monoclonal antibodies (MAbs). Such an isolation scheme involves multi-step procedures including ficoll-density fractionation and presort enrichment followed by cell sorting. Over the past decade, various cell-surface and metabolic markers have been identified and used to isolate human HSCs and progenitors as summarized in Table 1 . Among them, CD34 has become the most critical cell-surface marker for positively selecting a rare cell population (1 ,2 ). Within the CD34+ cell population, the differential expression of Thy-1, CD38, and AC133 have been used to fractionate HSCs and progenitors. In order to subfractionate CD34+ cells by these markers, the cells can be further purified by flow cytometry. HSCs can be further enriched into a Thy-1+ (3 7 ), CD38-lo (8 10 ), Thy-1+ CD38-lo (11 ), or AC133+ (12 ,13 ) fraction of CD34+ cells.

        Table 1

        Commercially Available Cell-Surface and Metabolic Markers for Isolation of Human HSC and Progenitor Cells

        Marker

        Expression/Remark

        Fluorochrome conjugate recommended

        Reference

        Positive marker

        CD34

        Positive

        FITC, PE, APC, BIO

        1,2,33

        Thy-1

        Positive

        PE, BIO

        3,4

        AC133

        Positive

        PE

        12,34

        Negative/low marker

        CD38

        Negative /low

        FITC, PE, APC

        8,9

        HLA-DR a

        Negative to low

        FITC, PE

        35,36

        Mature lineage marker, Lin-

        CD2

        T-cell lineage

        FITC, PE, BIO

        3

        CD3

        T-cell lineage

        FITC, PE, APC, BIO

        3

        CD19

        B-cell lineage

        FITC, PE, APC, BIO

        3

        CD16

        NK-cell lineage

        FITC, PE, APC, BIO

        3

        CD14

        Myeloid lineage

        FITC, PE

        3

        CD15

        Myeloid lineage

        FITC, PE

        3

        Glycophorin A

        Erythroid lineage

        FITC, PE

        3

        2nd Step reagent

        Avidin/Streptavidin

        For BIO MAb

        FITC, PE, APC, TXRD, PharRed, Cy-chrome d

         

        Metabolic marker b

        Rhodamine 123 c

        Low

        Mitochondria-binding dye

        37,38

        Hoechst 33342 c

        Low

        DNA-binding dye

        39,40

        Pyronin Y

        Low

        RNA-binding dye

        39,40

        Propidium iodide

        Negative to low

        Dead-cell exclusion

         

        Abbreviations: FBM, fetal bone marrow; MPB, mobilized peripheral blood; ABM, adult bone marrow; HSC, hematopoietic stem cells; FITC, fluorescein; PE, phycoerythrin; APC, allophycocyanin; TXRD, Texas red; BIO, biotinylated. a FBM, MPB HSCs express HLA-DR (41,42). b To isolate quiescent HSC. c Substrates for p-glycoprotein, encoded by MDR-1. HSC possess high levels of p-glycoprotein efflux activity. d Recommended for single laser flow cytometry only, lineage marker positive and PI positive cells can be excluded simultaneously.

        ad image
        提问
        扫一扫
        丁香实验小程序二维码
        实验小助手
        丁香实验公众号二维码
        扫码领资料
        反馈
        TOP
        打开小程序