What is a flow leukemia panel?

Flow Leukemia Panel: A Critical Diagnostic Tool for Hematologic Malignancies

A flow leukemia panel is a specialized multiparameter flow cytometry test that analyzes cell surface and cytoplasmic antigens (CD markers) to diagnose, classify, and monitor leukemias and other hematologic malignancies.

Components and Methodology

Flow leukemia panels utilize multiparameter flow cytometry to simultaneously assess multiple cellular properties at the single-cell level. The technology works by:

1. Sample preparation: Typically using bone marrow aspirate or peripheral blood with sufficient blast cells
2. Cell labeling: Using fluorescent-tagged monoclonal antibodies that bind to specific cell surface or cytoplasmic antigens
3. Analysis: Cells pass through a laser beam one at a time, and their light scatter and fluorescence properties are measured

Essential Markers in Flow Leukemia Panels

According to European Myeloma Network guidelines, comprehensive flow panels should include:

For Basic Identification of Cell Populations:

• CD38, CD138, and CD45 should all be included in at least one tube for proper cell identification and enumeration 1
• Primary gating should be based on CD38 vs. CD138 expression

For Acute Leukemia Classification:

• Minimal panel: Must include CD19 and CD56 for detection of abnormal cells 1
• Preferred panel: Should also include CD20, CD117, CD28, and CD27 1

Clinical Applications

Flow leukemia panels serve several critical purposes:

1. Primary diagnosis and classification of leukemia types:

   • Distinguishing between acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL)
   • Further subtyping into B-ALL, T-ALL, or specific AML subtypes
   • Identifying minimally differentiated or ambiguous lineage leukemias

2. Detection of minimal residual disease (MRD) after treatment:

   • Assessing treatment efficacy
   • Predicting outcomes
   • Determining stringent complete remission 1

3. Differential diagnosis between neoplastic disorders and reactive conditions 1

Specific Leukemia Patterns

Different leukemia types show characteristic immunophenotypic patterns:

• AML: Often CD34+ (62%), HLA-DR+ (86%), and typically positive for myeloid markers CD13 and CD33 2
• Acute Promyelocytic Leukemia (APL): Typically CD34-/HLA-DR- (80% of cases) 2
• B-ALL: Usually TdT+, CD19+, CD20+ (in 24% of cases) 2
• T-ALL: Typically cytoplasmic CD3+, CD5+, CD7+ 2

Workflow and Sample Requirements

According to ASCO clinical practice guidelines:

• Flow cytometry should be performed on bone marrow aspirate when possible 1
• If bone marrow aspiration is not possible (due to dry tap or patient condition), peripheral blood with sufficient blasts can be used 1
• Manual differential count, flow cytometry, FISH, and next-generation sequencing can be performed on peripheral blood if it shows sufficient blasts 1

Advantages and Limitations

Advantages:

• Rapid analysis (results available within hours)
• High sensitivity for detecting small populations of abnormal cells
• Ability to simultaneously assess multiple markers
• Useful for identifying unusual variants like minimally differentiated AML (M0) and biphenotypic leukemias 2

Limitations:

• Requires viable cells and adequate sample quality
• Loss of architectural relationships in tissue samples
• May miss certain diagnoses like Hodgkin lymphoma 3
• Discrepancies between flow cytometry and morphology cell counts are primarily related to sample quality 1

Common Pitfalls and Caveats

1. Sample quality issues: Ensure marrow elements are present in follow-up samples, particularly normal plasma cells in MRD negative cases 1

2. Timing considerations: Samples should be processed promptly to maintain cell viability

3. Interpretation challenges: Some leukemias may show aberrant marker expression or lack typical markers:

   • Some B-ALL cases lack CD34 expression (29%) 2
   • A small percentage of ALL cases are TdT-negative (5/180 B-ALL cases) 2

4. Technical limitations: Flow cytometry results should always be correlated with morphology, cytochemistry, cytogenetics, and molecular studies for comprehensive diagnosis

Flow leukemia panels have become an essential component of the diagnostic workup for acute leukemias, providing critical information that guides treatment decisions and improves patient outcomes.