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Flow Cytometry > Granulocytic Normal Differentiation > Flashcards

Granulocytic Normal Differentiation Flashcards

1
Q

What is the immunophenotype of general hematopoietic progenitor cells?

A

(+)
- CD34, CD133, CD184 and HLA-DR

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2
Q

Maturation from myeloblast to promeylocyte is characterized by what?

A
  • loss of CD34 and HLA-DR
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3
Q

Maturation from promyelocyte to myelocyte is characterized by what?

A
  • loss of CD117
  • gain of CD11b and CD11c
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4
Q

When is CD64 expressed?

A
  • promyelocytes through metamyelocytes
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5
Q

In the myeloid cell lineage, how are CD13 and CD33 expressed? What is the pattern of CD13 expression?

A
  • both are expressed at all stages of maturation
  • CD13
    • brightly expressed by blasts and neutrophils
    • moderately expressed by promyelocytes
    • dimly expressed by metamyelocytes
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6
Q

What is the pattern of CD33 expression in myeloid cells?

A
  • CD33 shows slightly decreased expression as the myeloid cell precursors mature
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7
Q

What is the pattern of expression of CD45 in myeloid precursors?

A
  • granulocytic precursors express CD45 dimly
  • the intensity of expression increases in the final stages of differentiation
    *based on the SSC and the intensity of CD45 neutrophils which are the brightest for CD45 can be separated from promyelocytes, etc.
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8
Q

What is the immunophenotype of normal granulocyte/macrophage progenitors?

A
  • CD34++
    -CD117 -/+
    -HLA-DR +
  • CD38 +
  • TDT +/-
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9
Q

What is the immunophenotype of normal myeloblasts?

A
  • CD34+
  • CD38+
  • CD117 ++
  • TDT -/+
  • CD13+ (dim)
  • CD33+
  • HLA-DR ++
  • MPO +
  • CD4 +
    -SSC low
  • FSC variable
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10
Q

When does CD117 expression start in myeloblasts?

A
  • CD117 expression appears after CD34

Note: CD34 is expressed by all hematopoietic precursors, including early myeloblasts

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11
Q

What is the pattern of expression of CD13 and CD33 in myeloblasts?

A
  • CD13 expression occurs first and increases as cells mature until the myelocyte stage
  • CD33 expression occurs after CD13
  • both markers increase in intensity in the myeloblasts
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12
Q

What is the immunophenotype of normal promyelocytes?

A
  • CD34 -
  • CD38 +
  • CD117 +
  • CD13 +
  • CD33 ++
  • HLA-DR -
  • CD4 +
  • CD15 dim+/-
  • MPO ++
  • CD65+
  • SSC high
  • FSC high
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13
Q

What is the pattern of expression of CD34 and HLA-DR in promyelocytes?

A
  • they lose CD34 and HLA-DR
  • but they retain CD117
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14
Q

What is the pattern of expression of CD13 and CD33 in promyelocytes?

A
  • CD13+
  • CD33 bright +

Note: they also show dim expression of CD4 and start to acquire CD15

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15
Q

At what transition is CD117 expression lost?

A
  • transition from promyelocytes to myelocytes the CD117 expression is lost

Note: CD13 expression is also dim at this stage and then gradually increases

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16
Q

What is the normal immunophenotype for myelocytes?

A
  • CD4+/-
  • CD117 -
  • CD13 dim+
  • CD33 +
  • CD11b dim+/-
  • CD15 +
  • CD65 +
  • MPO+
  • SSC high
  • FSC high
17
Q

What is the pattern of expression of CD13 at the myelocyte stage?

A
  • CD13 diminishes at the myelocyte stage and then gradually increases
18
Q

What marker starts to get acquired at the myelocyte stage?

A
  • CD11b
  • the intensity increases as the cell matures to late myelocytes and metamyelocytes

Note: while 11b expression is increases there is a loss of CD4

19
Q

What is the pattern of expression of CD33 in myelocytes?

A
  • CD33 progressively decreases
20
Q

What is the normal immunophenotype in metamyelocytes?

A
  • CD4-
  • CD13+
  • CD33+
  • CD35 dim+
  • CD11b+
  • CD15+
  • CD16+
  • CD65+
  • MPO+
  • SSC high
  • FSC high
21
Q

What markers begin to be expressed in metamyelocytes?

A
  • start to express CD10 and CD16 which increase in intensity as the cells progress to mature neutrophils
22
Q

What other markers are expressed at the metamyelocyte/band/segmented stages?

A
  • CD24
  • CD32
  • CD35
  • CD65
23
Q

What is the normal immunophenotype of segmented forms (bands/neutrophils)?

A
  • CD10++
  • CD13+
  • CD33+
  • CD35+
  • CD11b++
  • CD11c++
  • CD15++
  • CD16++
  • CD65+
  • MPO+
  • SSC high
  • FSC high
24
Q

What markers are unregulated in infections and in patients receiving G-CSF therapy?

A
  • CD14 and CD64
25
Q

What are the patterns of expression of the GPI anchored proteins on the myeloid lineage?

A
  • CD55 and CD59 are expressed constantly at all the stages of differentiation
  • CD59 shows no changes in the intensity of the expression
  • CD55 decreases from early precursors to promyelocytes and then increases in intensity to neutrophils
26
Q

In what situations can you see abnormal maturation patterns in the myeloid lineage?

A
  • MDS
  • BM regeneration after immunosuppressive therapy
  • toxic marrow insult (infections, nutritional deficiency, drugs)
  • increased hematopoietic cell turnover from peripheral destruction
  • uncontrolled expansion of neoplastic cells
27
Q

What are some clues to a possible false positive result?

A
  • may be caused by an old sample (increased SSC of neutrophils, decreased numbers of monocytes)
  • presence of a PNH clone
28
Q

What may cause a false negative result?

A
  • sample hemodilution
29
Q

How can EDTA samples alter immunophenotype expression?

A
  • known to influence marker expression
    *especially CD11b
  • some authors recommend the use of Heparin
30
Q

Where are basophils located on the CD45 vs. SSC plots?

A
  • located between blasts, hematogones and lymphocytes
31
Q

What is the immunophenotype that can differentiate them from neutrophils?

A
  • negative for CD15 and CD16
  • dimmer expression of CD13 compared to neutrophils
32
Q

What markers are basophils positive for?

A

(+)
- CD9, CD13, CD22, CD25(dim), CD33, CD36, CD38 (bright), CD123 (bright), CD45 somewhat dim

33
Q

What markers are basophils negative for?

A

(-)
- CD19, CD34, CD64, CD117, and HLA-DR

34
Q

Where are eosinophils found on SSC vs. FSC?

A
  • very low forward scatter and high side scatter
35
Q

How can eosinophils be differentiated from neutrophils?

A
  • dimmer expression of CD13 and CD15 (compared to neutrophils)
  • lack CD10 and CD16
36
Q

What is the immunophenotype of eosinophils?

A

(+)
- CD11b, CD11c, CD13, CD15, CD33, CD38

(-)
- CD4, CD10, CD16, CD34, CD117 and HLA-DR

Flow Cytometry (17 decks)

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