Population Identification (small Flow Book) Flashcards
What is the pattern of CD45 expression in hematopoietic cells?
- increasing intensity is seen on most maturing hematopoietic cells.
CD123 is a marker most helpful in identifying which populations?
- Basophils
- Plasmacytoid dendritic cells
What cell types will have the highest level of expression of CD45 ?
- mature lymphocytes
- monocytes
- granulocytes
Note: immature granulocytes usually have lower CD45 expression
What is the immunophenotype of the earliest precursor cells within the marrow?
- cells are capable of differentiating along a variety of lineages
- CD34(++), CD38 (low to negative), CD45, HLA-DR (intermediate)
- CD13 (intermediate), CD33 (low), CD117 (low), and CD123 (low)
What markers do blasts acquire when they transition to the myeloid lineage ?
- CD13, CD15, and CD33
- Note:
- promyelocytes have a high level of expression of CD13, CD33, and CD38
* the level of CD33 and CD64 is lower than a mature monocyte
- promyelocytes have a high level of expression of CD13, CD33, and CD38
What is the pattern of CD13 and CD16 in maturing myeloids ?
- myeloids lose CD13
- then they acquire CD13 and CD16 together
- both these antigens are highly expressed on the neutrophils
What are the initial markers that define myeloid maturation from an early precursor?
- begins when the CD34+ precursor starts acquiring CD38 along with:
- higher levels of CD13, CD33, and CD117
- with a slight decrease in CD45
What antigens are lost when myeloblasts move into the promyelocyte stage?
- CD34 and HLA-DR are lost
- CD13 expression increases as CD15 is acquired
- IMPORTANT:
- Promyelocytes have levels of CD33 and CD64 that are the highest for myeloid cells
But should still be less than mature monocytes
- Promyelocytes have levels of CD33 and CD64 that are the highest for myeloid cells
What happens to antigen expression as cells move into the myelocyte stage?
- There is a decrease in SSC, CD13 and CD33
What myeloid stage are CD13 and CD16 acquired together?
- Metamyelocytes
- Bands and neutrophils show high levels of CD13 and CD16 with maximal CD45
What antigenic changes can be seen in neutrophils in reactive conditions?
- can upregulate CD64 or downregulate CD16
- marrow-regeneration can lead to acquisition of CD56
- Remember
- PNH causes a loss of CD16
What antigens are obtained during monocytic maturation from the stem cells?
- begins in a similar fashion compared to myeloid maturation
- CD34+ acquire CD38
- then there are higher levels of CD13, CD33 and CD117
What markers indicate commitment to the monocytic lineage ?
- high levels of HLA-DR, CD11b, CD33, and CD64
* the expression of these antigens is much higher compared to myeloid precursors
What markers do immature monocytes demonstrate (monoblasts and promonocytes) ?
- they acquire CD15 with low variable expression of CD13
- Note:
- with progressive maturation (mature monocytes) express CD13, CD14 and CD45 at high levels
What is the activated monocyte phenotype?
- CD16+
- slightly decreased levels of CD14, CD33, CD64, and HLA-DR
- Note:
- CD14 can be lost in PNH monocytes
How is CD36 used in identifying monocytes ?
- CD36 is another marker that is acquired by promonocytes
- the issue is that platelets and erythroids can be positive for this as well
- issue becomes when platelets stick to things then you have non-specific expression
- also platelets can compete for the anti-CD36 antibody and decrease the intensity on monocytes
What antigens are acquired when the stem cell progenitor becomes erythroid?
- CD34+ progenitor acquires CD38
- high levels of CD117 are then acquired
What is antigen expression as the erythroid cells mature ?
- CD13 and CD33 are dropped (sometimes low levels are retained)
- then after that CD38, CD117 and CD45 are lost
- then CD71 is gained at a high level
- following this glycophorin and heme are acquired
When is the expression of CD71 lost in the erythroid precursor ?
- when the nucleus is lost, then CD71 is no longer expressed
- mature red cells are lysed by ammonium chloride for flow cytometry so evaluation of them is limited
How are eosinophils identified by flow cytometry ?
- gate around the high CD45 and SSC
- then subgate the negative CD16 population [eosinophils]
Where in the CD45 vs. SSC are basophils and PDCs found?
- “blast gate”
- they are differentiated from blasts by their expression of CD123
How are basophils and PDC’s differentiated by flow cytometry ?
- Basophils : negative for HLA-DR and positive for CD38 (bright)
- PDC: express high levels of HLA-DR
Where does B cell development start and what antigens are expressed/lost initially ?
- begins in the bone marrow
- stem cell (CD34+, CD38-) acquires CD38 and then loses CD13, CD117, and CD123
- then CD33 is lost after
What are the earliest markers expressed by B cell progenitors ?
- CD34, CD79a, TdT and CD10
- they then acquire CD19
- these progenitors are also considered to be hematogones
- hematogones show an inverse relationship with age
- they can be increased in regenerative marrows, immune mediated cytogenetic, and
Acquired immune deficiency
What are the three stages of hematogone maturation ?
- Stage I:
* CD19, TdT, CD34 and CD10 (bright) - Stage II:
- decreased CD10, loss of TdT and gradual loss of CD34
- Stage III:
- acquisition of CD20 with still some CD10 expression
Loss of CD10 completely means the mature B cell stage has been reached.
Where can naive mature B cells be found?
- all compartments (bone marrow, peripheral blood, tissues)
- primary follicles (predominantly), mantle zones of reactive follicles
What immunophenotypic changes are seen in B cells when they enter the germinal center and get exposed to antigen?
- Acquire CD10 and have increased CD38
- bright CD20 expression
When are CD5 positive, normal B cells encountered?
- these are called transitional B cells (B1)
- they are a type of naive B cells that expresses CD5
- more common in children and young adults but can be seen in adults
- they can be seen in all types of tissues, but frequently in head and neck
