flow


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  • Approach

    • B cells (CD19+)
      • K/L abnormal if monotypic or lack both
      • Aberrant: CD5, CD10
      • Distractors: CD10 on hematogones
    • T cells (CD3+)
      • Aberrant losses: CD3 (smear), CD7
      • CD4/8 coexpression
      • LGLs: CD56
      • CD4:8 ratio - normally 2 in blood, 0.5 in marrow
    • blasts (blast gate)
      • blast gate should be ~empty in blood, < 5% in marrow
      • CD34 should be ~0% in blood, < 5% in marrow
    • monos (mono gate)
      • aberrant CD56

  • Technique

    • detects CD markers on cells
    • # color flow = simultaneously detect # markers per tube
    • fluorochrome = molecule that emits the color

  • Tube combinations

    • every tube needs a CD45
    • acute screen
      • 34, 117, 56
    • B cell
      • 19, 5, 10
      • 20, 23, FMC7
      • 19+20, K/L
      • Hairy cell: 103, 25, 20, 11c, 22
    • T
      • 3, 7, 56
      • T: 4, 8, 38
      • LGL: 3, 56, 57
      • adult T cell: 3, 7, 25
      • ddx thymoma: CD1a, CD99, TdT
    • plasma cell (poor sensitivity by flow)
      • 138, K/L
      • 56, 19, 117
      • 38, 20, 3
    • myeloid
      • 16, 13, HLADR
    • specific lymphomas

  • Gating

    • FSC vs SSC: gates out debris (low FSC)
    • CD45 vs SSC: standard populations
    • CD45 vs FSC: subgate larger cells
    • specific CD markers

  • Cutoffs

    • negative: < 10 or same as neg ctrl
    • dim: < 10^2 or 1 log neg ctrl
    • moderate: < 10^3 or 1-2 logs neg ctrl
    • bright: > 10^3 or 2 logs neg ctrl

  • Normal markers

    • lymphocyte markers
    • myeloid markers
    • erythroid markers

  • PNH

    • gate granulocytes or monocytes
    • PNH clones have decreased GPI proteins CD55, 59

  • Signout

    • Normal: No monoclonal B cell, aberrant T cell, or expanded blast population identified
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