What is the appropriate workup to rule out lymphoma?

Workup to Rule Out Lymphoma

The diagnostic workup for suspected lymphoma must begin with an excisional lymph node biopsy to establish a definitive pathologic diagnosis, accompanied by comprehensive laboratory testing, contrast-enhanced CT imaging of the chest/abdomen/pelvis, and PET-CT for staging FDG-avid disease. 1, 2

Tissue Diagnosis: The Critical First Step

Excisional lymph node biopsy is the gold standard and must be performed immediately to obtain adequate tissue architecture for histologic evaluation, immunophenotyping, flow cytometry, and molecular studies required for accurate WHO classification. 1, 3, 4, 5

Biopsy Selection Hierarchy:

• First choice: Complete excisional biopsy of an entire lymph node 1, 3
• Second choice: Incisional biopsy (when excision would cause significant morbidity due to deep anatomic location or very large mass size) obtaining at least 1-2 cm³ of tissue 1
• Inadequate: Fine-needle aspiration alone is insufficient for initial diagnosis except in rare circumstances when combined with immunohistochemistry and judged diagnostic by an expert hematopathologist 1, 3, 5
• Discouraged: Core needle biopsy should not be used as the initial diagnostic method unless excisional biopsy is unsafe or impossible 1

Critical Specimen Handling:

• Tissue must arrive fresh and intact—never immerse in fixative before initial processing 1
• Section at 2 mm intervals perpendicular to the long axis 1
• Divide specimen for multiple fixatives: formalin for routine histology, B5/B+ for cytologic detail, and snap-frozen portions for immunohistochemistry and molecular work-up 1

Essential Laboratory Testing

Obtain the following blood tests immediately: 1, 2

• Complete blood count with differential and platelets to evaluate for bone marrow involvement, cytopenias, and abnormal lymphocyte populations 1, 2
• Lactate dehydrogenase (LDH) as a critical prognostic marker and component of the International Prognostic Index 1, 2
• Comprehensive metabolic panel including liver and renal function tests to assess organ involvement 1, 2
• β2-microglobulin for prognostic assessment 2
• Protein electrophoresis and serum/urine immunofixation 2
• Hepatitis B surface antigen and core antibody, hepatitis C antibody (mandatory before initiating anti-CD20 therapy due to reactivation risk) 1, 2
• HIV serology 2

Additional Testing for Specific Presentations:

• Flow cytometry of peripheral blood is mandatory in nodal marginal zone lymphoma and splenic marginal zone lymphoma, optional in extranodal marginal zone lymphoma 2
• Direct antiglobulin (Coombs) test optional in splenic marginal zone lymphoma 2
• Cryoglobulins and cryocrit if hepatitis C positive 2

Pathologic and Immunophenotyping Studies

The biopsy specimen must undergo comprehensive analysis: 1

For Hodgkin Lymphoma Suspicion:

• Immunohistochemistry panel: CD3, CD15, CD20, CD30, CD45, CD79a, PAX5 1

For Non-Hodgkin Lymphoma Suspicion:

• Pan-B cell markers: CD19, CD20, CD22 1
• Pan-T cell markers: CD3, CD5 1
• Additional markers: CD10, BCL2, BCL6, cyclin D1, Ki-67 1
• Flow cytometry for detailed immunophenotyping and clonality determination 1, 2
• Molecular testing for antigen-receptor gene rearrangements and BCL2 abnormalities 1
• Cytogenetic/FISH studies for t(14;18) and BCL6 rearrangements 1

Special Consideration for Marginal Zone Lymphomas:

• MYD88 gene mutation testing helps differentiate marginal zone lymphoma (rarely harbors MYD88 mutations) from lymphoplasmacytic lymphoma (MYD88 mutations in the large majority) 2

Imaging Studies

Mandatory Imaging:

• Contrast-enhanced CT of chest, abdomen, and pelvis (with oral and IV contrast unless contraindicated by renal insufficiency) 1, 2
• PET-CT from skull base to mid-thigh is the gold standard for staging FDG-avid lymphomas (most Hodgkin and aggressive non-Hodgkin lymphomas) and should be obtained no longer than 1 month before starting therapy 1, 3
• Chest radiograph 3

Site-Specific Imaging:

• CT or MRI imaging of orbits and salivary glands for marginal zone lymphomas 2
• Endoscopic ultrasound for gastric marginal zone lymphoma to define gastric wall infiltration and perigastric lymph node involvement 2
• Head CT or brain MRI if neurologic symptoms present 2

Important Nuance on PET-CT:

PET scanning was historically considered of little clinical utility in marginal zone lymphomas, but this is being reconsidered with increased sensitivity of modern PET-CT equipment. 2 It should be considered when only localized treatment is planned or when clinical/laboratory data suggest transformation to high-grade histology. 2

Bone Marrow Evaluation

Bone marrow biopsy is NOT routinely required in newly diagnosed Hodgkin lymphoma undergoing pretreatment staging with PET-CT if the PET scan is negative or displays homogeneous bone marrow uptake. 1

When Bone Marrow Biopsy IS Required:

• Mandatory in nodal marginal zone lymphoma and splenic marginal zone lymphoma 2
• Highly recommended in extranodal marginal zone lymphoma, particularly non-gastric lymphoma and when only local treatment is planned 2
• Essential for most non-Hodgkin lymphomas when treatment decisions depend on precise staging 2, 1
• Required if cytopenias are present despite negative PET findings 1

Bone marrow aspirate should include morphology and flow cytometry. 2

Clinical Assessment

Document the following during physical examination: 1, 3, 2

• All accessible lymph node regions including cervical, supraclavicular, axillary, inguinal, and Waldeyer's ring 3, 2
• Liver and spleen size 3, 2
• Performance status 2
• B symptoms: unexplained fever >38°C, drenching night sweats, or weight loss >10% within 6 months 1, 3
• Additional symptoms: fatigue, pruritus, alcohol-induced pain 3
• Testicular examination in males (testicular involvement rare but slightly more common in T-cell lymphoma) 2

Critical Pitfall:

Physical examination alone is inadequate for accurate lymph node assessment. Studies show physical examination underestimates lymph node size in all regions, with particularly poor correlation in supraclavicular (R² = 0.529) and axillary regions (R² = 0.368). 6 Always confirm with imaging.

Pre-Treatment Organ Function Assessment

Before initiating therapy, obtain: 1, 7

• Echocardiogram or MUGA scan to assess left ventricular ejection fraction when anthracycline-containing regimens are planned 1, 2
• Pulmonary function tests with DLCO when bleomycin-containing regimens (ABVD or BEACOPP) are planned 1, 7
• Pregnancy testing mandatory for women of childbearing potential 1, 2
• Fertility counseling should be offered before treatment initiation 1, 2

Special Considerations for Fever in Suspected Lymphoma

If fever is present, infection must be ruled out and treated first before attributing fever to lymphoma. 7

Immediate Fever Workup:

• Blood cultures (≥2 sets from separate sites) 7
• Urine culture 7
• Chest radiograph if pulmonary symptoms present 7
• Empiric broad-spectrum IV antibiotics immediately if neutropenic (ANC <500/mm³) without awaiting culture results 7

Lymphoma-related fever is a diagnosis of exclusion only after exhaustive infection workup is negative. 7

Staging Classification

Use the Lugano classification system which incorporates the Ann Arbor staging with modern PET-CT findings: 3, 4

• Stage I: Single lymph node region or single extralymphatic organ
• Stage II: Two or more lymph node regions on same side of diaphragm
• Stage III: Lymph node regions on both sides of diaphragm
• Stage IV: Diffuse or disseminated involvement of extralymphatic organs

Document presence of bulky disease (mass >10 cm or mediastinal mass >1/3 thoracic diameter) and B symptoms. 3

Common Pitfalls to Avoid

• Never rely on fine-needle aspiration alone for initial diagnosis—lack of architectural information leads to missed or incorrect classification 1, 3
• Never obtain inadequate tissue—this compromises accurate classification and treatment planning 1
• Never delay excisional biopsy in favor of less invasive procedures when lymphoma is suspected 1, 4
• Never attribute fever to lymphoma without first ruling out infection 7
• Never skip hepatitis B/C testing before anti-CD20 therapy—reactivation can be fatal 1, 2
• Do not perform bone marrow biopsy in Hodgkin lymphoma if PET-CT is negative for marrow involvement 1