Chronic Lymphocytic Leukemia - CLL

Last Literature Review: June 2023 Last Update:

Chronic lymphocytic leukemia (CLL) and small lymphocytic lymphoma (SLL) are different forms of a common malignancy that is characterized by the presence of small, mature neoplastic lymphocytes (referred to as monoclonal B lymphocytes) in the blood, bone marrow, and lymphoid tissues.    The diagnosis of CLL is usually made from blood cell counts and examination of peripheral smears. Additional laboratory testing, including cytogenetic testing, molecular testing, and flow cytometry, is used in prognosis and treatment decision-making. Determining the prognosis and appropriate treatment are especially important because CLL, which is usually indolent, may transform into a more aggressive disease such as diffuse large B-cell lymphoma (DLBCL) or Hodgkin lymphoma.

Quick Answers for Clinicians

How are chronic lymphocytic leukemia and small lymphocytic lymphoma diagnosed?

A CBC followed by peripheral smear interpretation and flow cytometry testing are generally sufficient to establish the diagnosis of chronic lymphocytic leukemia (CLL).  Evidence of lymphadenopathy by physical exam and/or imaging followed by a lymph node biopsy are necessary for the diagnosis of small lymphocytic lymphoma (SLL). 

Genetic tests, including gene sequencing, cytogenetics, and karyotyping, are used for prognosis and treatment decision-making. Bone marrow biopsy is not always necessary for diagnosis and prognosis because it is possible to test circulating lymphocytes for prognostic markers; however, biopsy may be helpful if there are concerns about immune-mediated disease or cytopenias that need to be evaluated before treatment. 

What role does next generation sequencing play in the investigation of chronic lymphocytic leukemia?

Next generation sequencing (NGS) can be used to assess for single gene variants, including substitutions and small insertions or deletions, that may have implications for prognosis or clinical management decisions. This testing complements the recommended immunophenotyping and cytogenetic workup of chronic lymphocytic leukemia (CLL) and other lymphoid malignancies and is used to tailor treatment (eg, in clinical trials), improve prognostication, and look for resistance mutations at relapse.

Which laboratory tests can be used to distinguish chronic lymphocytic leukemia from mantle cell lymphoma?

Several tests can distinguish between chronic lymphocytic leukemia (CLL) and mantle cell lymphoma (MCL), which is important for prognosis and treatment planning; such testing should always be performed at diagnosis. Unlike CLL, MCL exhibits the IGH-CCND1 fusion, t(11;14). MCL can often be excluded with flow cytometry testing for cyclin D1 [a surrogate marker for t(11;14)], or by using the gold standards of immunohistochemistry for cyclin D1 or fluorescence in situ hybridization (FISH) testing for IGH-CCND1.  Other tests that may exclude MCL include flow cytometry for CD200 and immunohistochemistry for LEF1 and SOX11.  MCL is usually negative for both CD200 and LEF1.  

Indications for Testing

Testing for CLL should be considered in adult individuals who present with abnormal CBCs (showing persistent, unexplained lymphocytosis lasting ≥3 months) and/or lymphadenopathy.

Laboratory Testing

Initial Evaluation

The initial evaluation for CLL/SLL should include a complete assessment of performance status and systemic symptoms, patient history, and a physical examination with an evaluation of the size of the liver and spleen.   Recommended laboratory tests include a CBC with differential and a metabolic panel.  Additional laboratory tests that may be useful include beta-2 (β2)-microglobulin, direct antiglobulin (direct Coombs), haptoglobin, lactate dehydrogenase (LDH), quantitative immunoglobulin (Ig), and uric acid tests.  A bone marrow aspirate with biopsy is no longer recommended for diagnosis but may be helpful if other tests are not diagnostic or to diagnose immune-mediated disease or cytopenias.   


A hematopathology slide review and immunophenotyping via flow cytometry on peripheral blood to detect and determine the concentration of monoclonal B lymphocytes are necessary to diagnose CLL, SLL, or monoclonal B lymphocytosis.    Monoclonal B lymphocytes must be present for at least 3 months to support a diagnosis. 

Diagnostic Criteria for CLL, SLL, and Monoclonal B Lymphocytosis
DiagnosisMonoclonal B Lymphocytes per LiterOther Features
CLL≥5 x 109Other atypical lymphocytes may be present but must be <55% of total lymphocytes
SLLaMay be <5 x 109Lymphadenopathy and/or splenomegaly
Monoclonal B lymphocytosisb<5 x 109None present

aConfirm diagnosis with histopathology on a lymph node biopsy specimen.

bMonoclonal B lymphocytosis is not a malignancy but may progress to CLL.

Sources: NCCN, 2023 ; Eichhorst, 2021 ; Hallek, 2018 

Flow Cytometry

Flow cytometry should be used to confirm the clonality of B cells in both CLL and SLL, which have the same immunophenotype.   Immunophenotyping with CD5, CD10, CD19, CD20, CD23, CD200, and kappa/lambda cell surface markers is recommended.  Additional testing [eg, flow or immunohistochemistry for cyclin D1 or fluorescence in situ hybridization (FISH) testing for the IGH-CCND1 fusion t(11;14)] may be useful to exclude similar conditions such as mantle cell lymphoma (MCL). 

Flow cytometry is also useful for prognosis and treatment decision-making.


Although histology is insufficient and generally not necessary to diagnose CLL, it should be performed to confirm an SLL diagnosis and may be necessary if flow cytometry fails to establish a CLL diagnosis.   Cytology or histology testing is also useful to exclude large-cell transformation to DLBCL, although this is rare. A lymph node biopsy specimen is preferred, followed by a bone marrow aspirate if the biopsy is nondiagnostic.   Fine needle aspiration or core needle biopsy is usually insufficient but can be performed in conjunction with other techniques if a lymph node is not readily accessible. 

Staging, Prognosis, and Treatment Decision-Making

Laboratory tests play an important role in staging, prognosis, and treatment decision-making in CLL. The prognostic significance of specific markers depends on the patient and treatment regimen.  In addition to flow cytometry and standard blood tests, the National Comprehensive Cancer Network (NCCN) recommends CpG-stimulated karyotyping, FISH testing for del(17p), immunoglobulin heavy chain variable region (IGHV) gene testing (if not already performed), and TP53 testing before treatment.  Other tests may also be helpful.  

Molecular Genetic Testing

IGHV Sequencing

Somatic IGHV sequencing to determine mutation status is particularly useful for prognosis and treatment decision-making, and the NCCN recommends somatic IGHV sequencing rather than flow cytometry testing.  Unmutated IGHV (≤2% mutated) has adverse prognostic implications, as do VH3-21 IGHV rearrangements, regardless of mutation status.  IGHV testing is only required once (ie, at diagnosis or before treatment) because IGHV mutation status does not change. 

Gene Panels by Next Generation Sequencing

Although genes with prognostic implications may be tested individually, next generation sequencing (NGS) is now preferred to evaluate many genes at one time. The following genes have implications for prognosis or clinical management in patients with CLL and may be included in such panels: ATM, BCL2, BIRC3, BTK, NOTCH1, PLCG2, POT1, and TP53. This is an emerging area of testing.

Flow Cytometry

Several flow cytometric markers have prognostic significance in CLL.  In cases in which it is not possible to test for IGHV mutation status, flow cytometry or other techniques to assess the expression of these markers may be considered instead, but this testing is not broadly recommended. 

Fluorescence In Situ Hybridization

In addition to recommended FISH testing for del(17p), FISH testing for del(11q), del(13q), and trisomy 12 is useful in prognosis and treatment decision-making.   The del(13q) abnormality has favorable prognostic implications in isolation, whereas both del(11q) and del(17q) have adverse prognostic implications. 


Identification of a complex karyotype by chromosome karyotyping analysis on CpG-stimulated CLL cells is useful in prognosis; a complex karyotype (≥3 unrelated abnormalities in multiple cells) has adverse prognostic implications.  Karyotyping is also useful in treatment decision-making for some patients, including those being considered for targeted treatment (eg, with Bruton’s tyrosine kinase inhibitor therapy). 

Other Testing

Testing for cytomegalovirus, hepatitis B, and hepatitis C is recommended both before and during treatment because some CLL treatments may reactivate these infections.   Pregnancy testing may also be useful before treatment. 

Monitoring for Treatment Response

Regular follow-up, including a CBC with differential, imaging, history, and physical examination, is recommended.  Examination of a bone marrow biopsy and minimal residual disease (MRD) testing may also be useful in some circumstances. 

Minimal Residual Disease

MRD testing is used in the context of clinical trials in CLL but is not currently recommended for monitoring in other contexts,    although it has potential for use in prognosis.  

ARUP Laboratory Tests


Flow Cytometry



Specimens: bone marrow, peripheral blood

For individual FISH probes, refer to the ARUP Oncology FISH Probes menu.

Cytogenomic Microarray

Specimens: bone marrow, peripheral blood

Molecular Testing

Polymerase Chain Reaction


For immunohistochemical tests that may be useful in the diagnosis or differential diagnosis of CLL, refer to ARUP’s Immunohistochemistry Stain Offerings.

Medical Experts



Picture of Nicole Leonard, MD
Nicole Leonard, MD
Anatomic and Clinical Pathology Resident, University of Utah School of Medicine and ARUP Laboratories


Bo Hong, MD, FACMG
Associate Professor of Pathology (Clinical), Co-Director of Laboratory Genetics and Genomics Fellowship, University of Utah
Medical Director, Cytogenetics and Genomic Microarray, ARUP Laboratories


Kristin Hunt Karner, MD
Associate Professor of Pathology (Clinical), University of Utah
Medical Director, Hematopathology and Molecular Oncology, ARUP Laboratories