How to Diagnose Chronic Lymphocytic Leukemia
The diagnosis of CLL requires ≥5,000 monoclonal B lymphocytes/µL in peripheral blood confirmed by flow cytometry showing the characteristic immunophenotype: CD5+, CD19+, CD20+ (dim), CD23+, with low surface immunoglobulin expression. 1, 2
Essential Diagnostic Criteria
The diagnosis is established when both of the following are present:
- ≥5,000 monoclonal B lymphocytes/µL in peripheral blood, sustained for at least 3 months 1, 3
- Flow cytometry confirmation of B-cell clonality with the characteristic immunophenotype 1, 2
Required Immunophenotype Pattern
CLL cells must demonstrate this specific pattern on flow cytometry 1, 2:
- Positive markers: CD5, CD19, CD23
- Dim/low expression: CD20, surface immunoglobulin, CD79b
- Light chain restriction: Either kappa OR lambda (not both)
Critical distinction: Mantle cell lymphoma also expresses CD5 but typically lacks CD23 expression; if CD23 is present, perform cyclin D1 staining or FISH for t(11;14) to exclude mantle cell lymphoma 1, 2
Diagnostic Algorithm
Step 1: Initial Laboratory Testing
When CLL is suspected, order:
- Complete blood count with differential showing absolute lymphocytosis ≥5,000/µL 1, 2
- Peripheral blood smear looking for small, mature lymphocytes with narrow cytoplasm, dense nucleus without nucleoli, and partially aggregated chromatin 1
- Smudge cells (ruptured lymphocytes appearing as nuclear shadows) are characteristic but not required for diagnosis 4
Step 2: Confirmatory Flow Cytometry
Flow cytometry of peripheral blood alone is sufficient for diagnosis—bone marrow biopsy is NOT required 1, 5
The flow cytometry panel must demonstrate 1, 2, 6:
- CD5+, CD19+, CD20+ (dim), CD23+
- Dim surface immunoglobulin (IgM or IgD)
- Dim CD79b
- Monoclonal light chain restriction (kappa or lambda)
- Additional markers: CD200+, CD43+ (bright), CD81+
Step 3: Pre-Treatment Evaluation
Once diagnosis is confirmed, perform these assessments before initiating therapy 1, 2:
Physical examination specifically evaluating:
- All lymph node regions (cervical, axillary, inguinal)
- Spleen size (palpation for splenomegaly)
- Liver size (palpation for hepatomegaly)
Laboratory tests:
- Serum chemistry including LDH, bilirubin
- Serum immunoglobulin levels
- Direct antiglobulin test (DAT) to detect autoimmune hemolytic anemia 1
Cytogenetic testing by FISH for del(17p) and del(11q)—this has direct therapeutic implications and should be performed before starting treatment 1, 2
Infectious disease screening (hepatitis B, hepatitis C, CMV, HIV) before chemoimmunotherapy or alemtuzumab to prevent viral reactivation 1
Step 4: Clinical Staging
Apply either the Binet (more common in Europe) or Rai staging system 1, 2:
Binet staging 1:
- Stage A: Hemoglobin ≥10 g/dL, platelets ≥100×10⁹/L, <3 lymph node regions involved
- Stage B: Hemoglobin ≥10 g/dL, platelets ≥100×10⁹/L, ≥3 lymph node regions involved
- Stage C: Hemoglobin <10 g/dL or platelets <100×10⁹/L (regardless of lymph node involvement)
Important: Clinical staging should be based on physical examination and blood counts, NOT on imaging studies 1
Common Pitfalls and Caveats
Bone marrow biopsy is NOT required for diagnosis but is recommended before initiating myelosuppressive therapy and for evaluating unexplained cytopenias 1
CT scans are NOT recommended for routine diagnostic workup outside of clinical trials 1
Monoclonal B-cell lymphocytosis (MBL) is diagnosed when monoclonal B lymphocytes are <5,000/µL without lymphadenopathy, organomegaly, or cytopenias; this progresses to CLL at 1-2% per year 2
Small lymphocytic lymphoma (SLL) is the same disease as CLL but with <5,000 B lymphocytes/µL in blood and presence of lymphadenopathy/splenomegaly; it requires the same immunophenotype and is treated identically 1, 5
The 3-month duration requirement for lymphocytosis ≥5,000/µL distinguishes CLL from transient reactive lymphocytosis 1, 3