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Thread: The Interpretation of Flow Cytometry

  1. #1

    Default The Interpretation of Flow Cytometry

    Gate CD45 and Side Scatter


    • Mature lymphocytes are characterized by low side scatter and strong CD45 expression.
    • Monocytes have higher SS and strong CD45 expression.
    • Erythropoietic precursors are CD45 negative and have low SS.
    • Granulopoietic precursors and granulocytes are weakly CD45 positive and have high SS.
    • Early hematopoietic precursors of various lineages, including CD34+ stem cells, are characterized by low CD45 expression and low SS.


    Granulocytic Differentiation

    • Precursors (promyelocytes): CD13+ (high levels), CD117+
    • Intermediate precursors (myelocytes): down-regulated dimly expressed CD13
    • Segmented neutrophils: up-regulated expressed CD13
    • CD11b and CD16 are initially expressed at low levels, but their expression increases during maturation.


    Monocytic Differentiation

    CD14, CD36, and CD64 are considered as monocyte-associated markers, CD14 being the most specific.

    During maturation toward promonocytes, progenitors down-regulate CD34 and CD117 and gain the expression of CD64, CD33, HLA-DR, CD36, and CD15, with an initial mild decrease in CD13 and an increase in CD45.

    Maturation toward mature monocytes leads to a progressive increase in CD14, CD11b, CD13, CD36, and CD45, with a mild decrease in HLA-DR and CD15.

    Mature monocytes show expression of bright CD14, bright CD33, variably bright CD13, bright CD36 and CD64, and low CD15.

    Erythropoietic Differentiation

    Early erythropoietic precursors are found in the blast area and can be identified by very bright CD44, bright CD71, intermediate CD36, positivity for HLA-DR, and expression of CD117 with "dim" CD45. Glycophorin A (CD235a) is expressed at a low level at this stage.

    Maturation to the basophilic erythroblast is accompanied by a decrease in CD44, disappearance of CD45 and acquisition of bright CD235a expression.

    At transition to the polychromatophilic/orthochromatophilic stage, erythroblasts show loss of HLA-DR, further decrease in CD44, and a mild decrease in CD36.

    Lymphocyte Differentiation

    B-cells

    • Early B (E-B) stage: CD34+, CD4+, terminal deoxynucleotidyl transferase (TdT)+, CD79a+, CD19-
    • Pro-B-cell stage: CD34+, CD19+, CD10+, TdT+, CD20-, cytIgM-
    • Pre-B: down-regulation of CD34 and TdT
    • Immature (IM)-B-cells: CD34-, CD19+, CD20-, CD10dim/-, IgM+
    • Mature B-cells: CD10-, CD19+, CD20+, IgM+, IgD+


    T-cells
    T-cell-restricted precursors: pre-alpha, CD34+, CD7+, CD45RA+
    Progenitors: CD34+, CD10+, CD24-
    Mature T-cells: CD7+, CD5+, CD2+, CD3, CD4+ or CD8+ (only one of the both)

    Immature Cells of Normal Bone Marrow

    CD34+ hematopoietic progenitor and precursor cells (HPC) that constitute most cells of the CD45/dim (blast) region are a heterogeneous cell population.

    A small fraction of pluripotent stem cells with long-term repopulating cell activity have been associated with the CD34/CD38- phenotype.

    CD34/CD45dim cells also include a major fraction of HPC already committed to different hematopoietic lineages.

    Human stem cells are defined by expression of CD90 and CD49f and are CD45RA negative.

    Early myeloid progenitors were isolated based on the expression of IL-3 receptor, a chain (CD123) or FLT3 (CD135) and CD45RA.
    Last edited by CheneyHsiung; Sun 12th June '16 at 5:30pm.
    Clinical Pharmacy Specialist - Hematology

  2. #2

    Default Examples of 10-Color Flow Cytometry Panels in Immunophenotyping of Leukemia and Lymphoma

    Examples of 10-Color Flow Cytometry Panels in Immunophenotyping of Leukemia and Lymphoma

    The Interpretation of Flow Cytometry-screen-shot-2016-06-12-at-4-43-10-pm-png
    Clinical Pharmacy Specialist - Hematology

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