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​Lab Guide
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Test ID: Flowcytometry Leukemia Lymphoma
Flowcytometry Leukemia Lymphoma
Useful For

Evaluating lymphocytosis of undetermined etiology.
Distinguishing acute lymphoblastic leukemia (ALL) from acute myeloid leukemia (AML).
Subtyping of Acute Lymphoblastic Leukemia according to cell lineage.

Subtyping of Acute Myeloid Leukemia according to cell lineage.
Distinguishing between lymphoma and acute leukemia.
Identifying B- and T-cell lymphoproliferative disorders involving bone marrow.
Phenotypic sub classification of B- and T-cell chronic lymphoproliferative disorders.
Recognizing monoclonal plasma cells for the diagnosis of plasma cell neoplasms.

Follow up and disease monitoring.

Assessment of disease prognosis.
Method name and description

Immunophenotyping by flow cytometry.

This test is based on the ability of specific antibodies to bind to the antigenic determinants expressed by cells. Surface staining of the Antigens is performed by incubating the sample with the Antibodies . The red cells are then removed by lysis and the sample is washed (Stain then Lyse technique). The sample is analyzed by flow cytometry analyzer. (Lyse then stain technique) may be used for some antigen’s determination. Flow cytometric immunophenotyping is performed using specific panels of antibodies:

Lymphoproliferative disorders:

Triage Panel: CD3, CD4, CD5, CD7, CD8, CD10, CD19, CD20, CD45 and kappa and lambda light chains.
If any further testing is required for diagnosis and subclassification, the additional panel may include:
-Markers for B-Cell Disorders: CD5, CD10, CD11c, CD19, CD20, CD22, CD23, CD25, CD38, CD43, CD45, CD49d, CD79b,CD103, CD200, BCL2, FMC7, IgD and IgM.
-Markers for T-Cell Disorders: CD2, CD3, CD4, CD5, CD7, CD8,CD25, CD26, CD45, cTIA,  TCR alpha/beta, and gamma/delta.

-Immature markers: Tdt, CD34 (as needed).
-Markers for Killer Disorders:: CD3, CD8, CD16, CD56 and CD57.


Acute  leukemia :

Panel: CD2, CD3, CD4, CD5, CD7, CD9, CD10, CD11b, CD11c, CD13,  CD14, CD15, CD19, CD20, CD33, CD34, CD36, CD45, CD56, CD64, CD117, HLA-DR, cytoplasmicCD3, cytoplasmic CD79a, cytoplasmic MPO and nuclear Tdt.


Possible Additional markers:
-B-cell ALL markers: CD19 , CD22 and IgM (surface & cytoplasmic).
-T-cell ALL markers: CD1a, CD2, CD3, CD4, CD5, CD7, CD8, TCR alpha/beta and gamma/delta.

-Others: CD41, CD61, CD71, GA and CD123.

Plasma Cell Neoplasm:

Panel: CD19,CD20,  CD38, CD45, CD56, CD117,  CD138, and cytoplasmic kappa and lambda light chains.
Clinical information

Diagnosis of Hematological Neoplasms is based on integrating morphologic, immunophenotyping, cytogenetic and molecular data.
Immunophenotyping can help resolve many differential diagnostic problems posed by the clinical or morphologic features.
Aliases

N/A
Specimen type / Specimen volume / Specimen container

Peripheral blood
Body fluids
Tissue specimens/FNA
** For Bone Marrow specimen, please order Flowcytometry Bone Marrow.

Peripheral blood in EDTA. At least 2 ml.

Body fluids (should be fresh) in plain container (no anticoagulant or additives) and for bloody specimens, please keep in EDTA tube. Quantity is dependent on specimen cellularity. Large volumes are preferred. For CSF, should be collected in sterile plain container.

Tissue specimens/FNA are placed in 15 mL plain container filled with ~ 5ml RPMI media.  Please send intact specimen (do not mince). Required tissue size at least (0.5 x 0.5 x 0.5) cm. Smaller tissue specimens may be acceptable but may limit the number of markers performed.
Tissue specimens should be fresh and sent ASAP to histopathology laboratory and the Histopathologist will assess if Flowcytometry indicated.

 
Collection instructions / Special Precautions / Timing of collection

For specimens received from outside HMC facilities should be accompanied by complete/proper patient identification on both the request form and the specimen container. The request form should state the clinical data and specify the order requested.

Regular working hours from Sunday to Thursday (7:00 AM-3:00 PM); however, Cutoff time of receiving specimens is 11:30 AM.

Specimens should be transported to the lab as soon as possible after collection during working hours of the laboratory.

** Tissue specimens placed in 15 mL plain container filled with ~ 5ml RPMI media should be fresh and sent ASAP to histopathology laboratory and the Histopathologist will assess if Flowcytometry is indicated


** Any Sample submitted to flow cytometry MUST NOT be put into FIXATIVE
Storage and transport instructions

4-28 ◦C
Specimen Rejection Criteria

Aged specimen (>24hour) with increased number of dead (non-viable) cell.

Frozen/ over-heated specimens.

Wrong specimen container.

Leaking or contaminated container.

Unlabeled or improperly labeled container.

Mislabeled specimen.

Improperly filled request form.

No specimen types.

Gross contamination of the specimen.

No request/electronic order received.
Biological reference intervals and clinical decision values

An interpretive report will be provided.
An interpretation of the immunophenotypic findings and correlation with the morphologic features will be provided by a hematopathologist for every case.
Turnaround time / Days and times test performed / Specimen retention time

4 working days. Preliminary report might be released within 8 working hours for acute leukemia cases.

Regular working hours from Sunday to Thursday (7:00 AM-3:00 PM); however, Cutoff time of receiving specimens is 11:30 AM.

* For requests after working hours and weekends, prior approval from the lab on-call pathologist by the attending consultant is mandatory for arrangements of the testing.

Specimen retention time: 7 Days.

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