1. Peripheral blood Lymphoma A panel cheatsheet
Jean Oak

2. Lymphoma A This is what you order for initial lymphoma assessment
What it is looking for in a nutshell
Mature B or T cell malignancy
Doesn’t assess blasts or myeloids
Tubes
B cell tubes
CD5/CD38/CD45/CD19
Lambda/kappa/CD45/CD19
CD20/CD10/CD45/CD19
FMC7/CD23/CD45/CD22
T and NK cell tubes
CD8/CD3/CD45/CD4
CD7/CD3/CD45/CD2
CD57/CD8/CD45/CD3
CD16/CD3/CD45/CD56

3. Goal – identify aberrant CD5+/CD19+ B cells, plasma cells, etc
Tube - CD5/CD38/CD45/CD19
Goal – identify aberrant CD5+/CD19+ B cells, plasma cells, etc
Look for aberrant
B cells
here1
Distinct population here is abnormal2
Likely
T cells
Normal B cells have a gradient
B cells are normally here
Plasma cells are usually here
Rare cells ok (late stage hematogones); CD5 in B cell lymphomas tend to be slightly dimmer than in T cells
Ddx: plasma cell/LPL (high CD38), hematogones, blasts, GC B cells

4. Kappa:Lambda ratio is usually 2-3:1
Tube - Lambda/kappa/CD45/CD19
Goal – Look for surface light chain restriction
B cells with l light chain
(these numbers should match)
Nonspecific
Monocytes with nonspecific binding
via Fc receptors
B cells with k light chain
(these numbers should match)
Kappa:Lambda ratio is usually 2-3:1
Suspect a clonal population if ratio >4:1 (kappa-restricted) or <1:1 (lambda-restricted)

5. Look for abnormal population here1
Tube – CD20/CD10/CD45/CD19
Goal – Look for B cells with aberrant CD10 expression or loss of CD20
Look for abnormal population here1
B cells are normally here
(numbers should match)
Abnormal1
PB B cells with dim/negative CD20 is abnormal2
Ddx: ALL/hematogones (likely CD20(-), CD45(dim), low FSC), post-GC derived lymphoma
Ddx: CLL, ALL, DLBCL

6. ~5% of T cells are CD4/CD8 double negative = normal gd T cells
Tube – CD8/CD3/CD45/CD4
Goal – Look for aberrant T cells
~5% of T cells are CD4/CD8 double negative = normal gd T cells
(Note % and are ~5%)
CD8 T cells
Probably NK cells
CD4 T cells
LGLs
CD4 T cells
Monocytes
(note dimmer CD4)
T cells are ~70-80% of peripheral blood lymphocytes
CD4:CD8 ratio is around 2-3:1

7. Usually naïve T cells predominate in the PB.
Tube – CD57/CD8/CD45/CD3
Goal – Look for aberrant CTL
Normal CD57
pattern
‘Memory‘ T cells NK
CD8+ T
CTL
(LGLs)
Mostly CD4+ T
NK, NKT
Naïve T cells
Usually naïve T cells predominate in the PB.

8. Normal T cells live here
Tube #6 – CD7/CD3/CD45/CD2
Goal – Look for aberrant T cell antigen loss
Normal T cells live here
Normal
gradient* NK NK
*Look for distinct populations – a gradual loss (“tail”) can be seen with aging or activation

9. FMC7 – Antibody against a different epitope of CD20
Tube #7 – FMC7/CD23/CD45/CD22 (aka CLL tube)
Goal – Rule in/out CLL/MCL. This is the best tube to calculate %B cells for MBL cases
HCL
CLL
HCL*
CLL
MCL
MCL
CLL
MCL
FMC7 – Antibody against a different epitope of CD20
*Dx of HCL will require additional testing (morphology, HCL extension tube)

10. Goal – NK/NKT cell characterization
Tube #8 – CD56/CD16/CD45/CD3
Goal – NK/NKT cell characterization
NKT
NKT NK NK

11. Goal – Check specimen viability/cell membrane permeability*
Tube #8 – CD45/7AAD
Goal – Check specimen viability/cell membrane permeability*
Live Dead
*Dead cells have permeable membranes so 7AAD leaks into the cell
Why we care: dead cells can have nonspecific binding of antibodies

12. 2) Correlate with calculated k:l ratio here
Re-gating of Lambda/kappa/CD45/CD19 for all CD19+ cells
Goal – Look at B cells as one group for light chain restriction
Gate of all B cells
1) Visually
check the k:l ratio
2) Correlate with calculated k:l ratio here
Light chain expression of all cells

13. Lymphoma A versus other lymphoma panels
Lacks
Additional tubes
Lymphoma B
Most T/NK cell tubes
CD7/CD3/CD45/CD2
CD57/CD8/CD45/CD3
CD16/CD3/CD45/CD56
CD16/CD13/CD45/CD56
Lymphoma C
FMC7/CD23/CD45/CD22 (CLL tube)
None
Cytopenia
FMC7/CD23/CD45/CD22
CD38/CD56/CD45/CD34 (blast tube)