Clinical Guide to Biomarkers for Non-Hodgkin Lymphoma

Non-Hodgkin lymphoma (NHL) is not one disease. It is a family reunion of many related cancers that arise from the immune system, each with its own personality and pace. Biomarkers are how we read that personality. They help answer the big questions: What subtype is this? How aggressive is it? What is the best way to monitor it?¹ If you have ever watched a fitness tracker turn steps into insight, biomarkers serve a similar role for lymphoma biology. They translate what is happening in the tumor into data that doctors can interpret.

This guide walks through the biomarkers that matter most in NHL, how they are used in real clinical workflows, how they influence diagnosis and monitoring, and where the limits are. The language is kept plain and precise medical terms are brought in only where they add clarity.

Why biomarkers matter in NHL

NHLs usually start in B cells or T/NK cells, the white blood cells that run immune surveillance. Under the microscope, many lymphomas can look similar. Biomarkers bring specificity. They fall into a few buckets:

• Diagnostic: Identify the exact lymphoma subtype and its cell of origin.¹
• Prognostic: Estimate risk of progression or relapse.²
• Predictive: Suggest whether a tumor is likely to respond to a targeted approach.
• Monitoring: Track response or detect minimal residual disease (MRD).³

Modern classifications, including the 5th edition WHO framework and International Consensus Classification, rely on a blend of tissue markers, chromosomal changes, and gene mutations.¹ ⁴ In practical terms, the right biomarkers help avoid mislabeling, reduce uncertainty, and align care with the tumor's actual biology. Large validation studies back many of these markers, though specific tests vary by center and region.

The diagnostic starting line: the biopsy

Everything starts with tissue. An excisional lymph node biopsy is ideal because it preserves architecture, which is crucial for classification.¹ When that is not possible, a core needle biopsy can still deliver enough for most studies. Fine-needle aspiration alone often falls short because it lacks tissue context.

From a single biopsy, a whole toolkit comes into play:

• Immunohistochemistry (IHC): Protein "badges" on cells, such as CD20 on B cells or CD3 on T cells.
• Flow cytometry: Sorting cells by their surface patterns and light-chain restriction to prove clonality.
• Cytogenetics and FISH: Chromosomal rearrangements; for example, t(14;18) in follicular lymphoma.
• Molecular testing: Targeted sequencing for mutations like MYD88 or EZH2.²
• EBER in situ hybridization: Detects Epstein–Barr virus in tumor cells where relevant.

Core biomarkers by common NHL subtype

Diffuse large B-cell lymphoma (DLBCL)

DLBCL is the most common aggressive NHL. It is a category with subtypes whose differences show up in biomarkers and outcomes.

Cell of origin:

• Gene-expression profiling divides DLBCL into germinal center B-cell (GCB) and activated B-cell (ABC) types.⁵
• When gene-expression testing is not available, the Hans IHC algorithm approximates this using CD10, BCL6, and MUM1.

Why it matters: ABC-type tumors tend to behave more aggressively than GCB in historical cohorts, a finding reproduced across studies.⁵ This is useful context when interpreting prognosis and considering targeted strategies in academic settings.

Genetic hits that raise risk:

• MYC rearrangements, especially combined with BCL2 and/or BCL6 rearrangements ("double-hit" or "triple-hit" high-grade B-cell lymphoma).⁶
• Dual protein overexpression of MYC and BCL2 by IHC ("double expressor") is a separate, less specific risk signal.

Serum and clinical markers:

• LDH reflects cellular turnover. Higher LDH often means higher tumor burden.²
• International Prognostic Index (IPI) uses age, stage, LDH, performance status, and extranodal disease to stratify risk, validated in thousands of patients.² ⁷

Monitoring biomarkers:

• Interim and end-of-therapy PET-CT, interpreted with the Deauville score, estimates metabolic response.⁸
• Circulating tumor DNA (ctDNA) assays can track tumor-specific DNA in blood.³ Multiple studies show ctDNA drops correlate with response and can presage relapse by months, though assays and thresholds vary.

Follicular lymphoma (FL)

FL is typically indolent but can transform into a more aggressive form, often resembling DLBCL. Key biomarkers:

• t(14;18)(q32;q21) involving BCL2; detected by FISH or PCR. Common but not universal.⁹
• EZH2 mutations (often at Y641) in a subset. These define a biologically coherent group and can inform research-driven strategies.
• FLIPI and FLIPI2 use clinical factors to estimate risk; they complement but do not replace biologic markers.⁹
• Beta-2 microglobulin correlates with tumor burden and prognosis; interpret carefully if kidney function is reduced.

MRD tracking using BCL2-IGH rearrangements has been studied, but routine use varies and is not universally adopted.³ PET-CT is helpful for staging and to evaluate suspected transformation when symptoms, LDH, or growth pattern change abruptly.

Mantle cell lymphoma (MCL)

MCL often carries t(11;14)(q13;q32) leading to cyclin D1 overexpression. If cyclin D1 is negative, SOX11 supports the diagnosis in most cases.

• TP53 mutations or 17p deletions identify a higher-risk subset with distinct behavior.¹⁰ ¹¹
• MIPI (Mantle Cell Lymphoma International Prognostic Index) incorporates age, performance status, LDH, and leukocyte count.¹⁰
• Proliferation by Ki-67 adds prognostic weight; higher indices signal more aggressive biology.¹⁰ ¹²

Marginal zone lymphomas (MZL), including MALT

These are usually indolent. Biomarkers help distinguish pathogenesis and guide expectations:

• Gastric MALT lymphoma can be driven by chronic Helicobacter pylori infection; EBER-negative by definition.
• t(11;18)(q21;q21) API2-MALT1 predicts resistance to antibiotic eradication strategies in gastric MALT and is a stable, helpful marker when present.
• Trisomy 3 and 18 are common but nonspecific.

In non-gastric MZL, infectious triggers vary by site (e.g., hepatitis C with some splenic MZL). Testing is tailored to the clinical picture.

Chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL)

CLL/SLL straddles hematology and lymphoma. It is rich in biomarker-guided risk tools with strong evidence:

• IGHV mutation status: Mutated IGHV generally signals more indolent disease; unmutated is higher risk.¹³ ¹⁴
• TP53 aberrations (del(17p) or TP53 mutation): Strong adverse prognostic impact across treatment eras.¹⁴ ¹⁵
• Additional cytogenetics: del(11q), trisomy 12, del(13q). Each carries distinct risk implications.
• CLL-IPI combines age, stage (Rai/Binet), beta-2 microglobulin, IGHV, and TP53 status.¹³
• MRD: High-sensitivity flow cytometry or next-generation sequencing can quantify residual disease down to very low levels. This is one of the best-studied MRD use cases in lymphoma-related disorders.³

Peripheral blood is often sufficient for diagnosis and monitoring in CLL, which makes biomarker follow-up more straightforward than in nodal-predominant lymphomas.

Burkitt lymphoma

This is a highly aggressive B-cell lymphoma with fast doubling times. Biomarkers are crisp:

• MYC translocation, typically t(8;14) with IGH, or variants with IGK/IGL.¹⁶ ¹⁷
• Very high proliferation index (Ki-67 near 100%).
• EBV positivity in most endemic cases and in a subset of sporadic cases.

Because growth is so brisk, labs that reflect turnover such as LDH and uric acid are often elevated, which is relevant for anticipating tumor lysis risk.

T-cell and NK-cell lymphomas

These are diverse, and biomarkers are essential for classification:

• Anaplastic large cell lymphoma (ALCL): ALK-positive versus ALK-negative status shapes classification and prognosis.¹⁸ DUSP22 and TP63 rearrangements further refine ALK-negative disease.
• Peripheral T-cell lymphoma, NOS: IHC panels, T-cell receptor (TCR) clonality, and gene expression studies support diagnosis but are less standardized than in B-cell disease.
• Cutaneous T-cell lymphoma (e.g., Sézary syndrome): Flow cytometry shows aberrant T-helper phenotype (CD4-positive with loss of CD7 or CD26) and a dominant TCR clone. Blood involvement is graded by tumor burden.
• Extranodal NK/T-cell lymphoma, nasal type: Uniformly EBV-positive by EBER, which is a defining feature.

Waldenström macroglobulinemia/lymphoplasmacytic lymphoma

Here, the biology blends B-cell lymphoma with plasma-cell features. Core markers:

• MYD88 L265P mutation in the majority of cases; highly informative but not exclusive.¹⁹ ²⁰
• CXCR4 mutations in a subset; can influence disease behavior.¹⁹
• Quantitative IgM level and serum viscosity relate to symptom risk. Free light chains add context when paraproteins are complex.

Blood tests you will actually see on a lab report

Not every useful biomarker requires specialized sequencing. Several common labs offer real signal:

• LDH: A surrogate for cell turnover. Trends matter more than a single value. Strenuous exercise or hemolysis can transiently raise LDH, so context is key.²
• Beta-2 microglobulin: Reflects tumor burden and renal handling. Elevated levels are prognostic in several subtypes but can rise with kidney impairment.
• Complete blood count (CBC): Anemia, neutropenia, thrombocytopenia, or lymphocytosis signal marrow involvement or immune disruption.
• Uric acid and phosphorus: Indirect markers of rapid cell breakdown and risk for tumor lysis physiology.
• Hepatitis B/C and HIV serology: Not lymphoma biomarkers per se, but essential context because they influence biology, risk, and safe treatment planning.

In select T/NK-cell lymphomas, quantitative EBV PCR in blood can parallel disease activity, though false positives from incidental reactivation can occur.

Imaging as a functional biomarker

Fluorodeoxyglucose PET-CT lights up metabolically active tissue. For most aggressive B-cell lymphomas, PET-CT at baseline and after therapy provides a strong readout of response using the Deauville scoring system. A complete metabolic response on end-of-therapy PET correlates with favorable outcomes in large cohorts.⁸ Pitfalls include uptake from infection or inflammation; a tooth abscess or recent vaccine can glow and confuse the picture. Some indolent lymphomas have lower or variable FDG avidity, so CT imaging and clinical context carry more weight there.

Circulating tumor DNA and minimal residual disease

ctDNA turns tumor biology into a blood test. There are two big approaches:

• Tumor-informed assays: Sequence the tumor to identify mutations or rearrangements, then build a personalized panel to detect those signals in blood.³
• Tumor-naïve panels: Use predefined lymphoma hotspots to scan blood directly, trading sensitivity for speed and simplicity.

What the evidence shows: In DLBCL and primary mediastinal large B-cell lymphoma, early ctDNA clearance aligns with better outcomes, and rising ctDNA can precede imaging relapse by months.³ In follicular lymphoma, ctDNA dynamics mirror tumor burden and may flag transformation risk when paired with clinical changes. In CLL, MRD measurement by high-sensitivity flow cytometry or NGS is particularly mature — depth of remission strongly correlates with duration of control in multiple trials.

Limitations to remember:

• Not all tumors shed DNA equally; deep lymph nodes can be underrepresented in blood.
• Clonal hematopoiesis (age-related mutations in blood cells, often DNMT3A/TET2/ASXL1) can create noise if not filtered by bioinformatics.
• Assay sensitivity and turnaround times vary by platform and laboratory.

Risk tools that synthesize biomarkers

Doctors often blend lab and clinical data into validated risk scores. These do not replace judgment but offer an evidence-based baseline for discussion.

• IPI, R-IPI, and NCCN-IPI in DLBCL: Age, stage, performance status, LDH, and extranodal sites combine to estimate risk. They were built and validated in large datasets and still anchor trial design.² ⁷
• FLIPI/FLIPI2 in follicular lymphoma: Age, stage, hemoglobin, nodal areas, LDH, and beta-2 microglobulin form the backbone.⁹
• MIPI in mantle cell lymphoma: Adds leukocyte count to age, performance status, and LDH.¹⁰
• CLL-IPI: Fuses clinical stage with IGHV, TP53, and beta-2 microglobulin for a powerful stratifier.¹³

Think of these like the composite health scores in your smartwatch. No single metric tells the whole story. The strength is in the combination.

Biomarkers that steer targeted decisions

Some markers connect directly to a target on the lymphoma cell. A few examples help anchor the concept:

• CD20 expression in most mature B-cell lymphomas supports use of CD20-directed strategies; lack of CD20 redirects thinking.
• CD19 expression is relevant for cellular therapies that look for this marker to find and eliminate B cells.
• CD30 expression in ALCL and in a subset of large B-cell lymphomas can be leveraged in specialized contexts.¹⁸
• ALK positivity defines a biologically distinct ALCL with different expectations than ALK-negative cases.¹⁸
• PD-L1/PD-L2 gains in primary mediastinal large B-cell lymphoma co-travel with 9p24.1 alterations and inform immunotherapy discussions in appropriate settings.

The unifying theme is fit: the biomarker has to match the biology of the treatment. This is where a precise pathology report pays dividends.

Life stage, sex, and special situations

Age is the most consistent modifier across NHL. Incidence rises with age, as does the prevalence of clonal hematopoiesis that can muddy ctDNA interpretation. Frailty influences how far one can chase a diagnostic workup in a single sitting, and which monitoring tests make sense.

Pregnancy requires a different playbook. Biopsy remains feasible, but imaging leans on ultrasound and MRI. PET-CT is generally deferred. Blood-based monitoring becomes more attractive where validated, though data are limited in pregnancy. Multidisciplinary planning is standard in these cases.

Pediatric and young adult NHLs include entities with unique biology, such as ALK-positive ALCL and Burkitt lymphoma. These are best navigated at centers experienced in adolescent and young adult oncology, where the biomarker panels and response criteria are age-tailored.

Practical pitfalls and caveats

Every test has an Achilles' heel. A few common ones to keep in mind:

• IHC variability: Antigen retrieval and fixation times can shift staining intensity. Reading context and controls matters.
• Small biopsies: Limited tissue can constrain the panel and make subtyping harder. Re-biopsy can be necessary if results and the clinical picture diverge.
• Steroids before biopsy: Brief symptom relief can come at the cost of blunted tumor morphology. If possible, avoid steroids until tissue is secured.
• LDH and beta-2 microglobulin: Exercise, hemolysis, and renal function move these numbers. Trend plus context beats a single spike.²
• EBV PCR: Low-level viremia is common and not specific. EBER positivity in tumor tissue is the more definitive link in EBV-driven lymphomas.
• ctDNA noise: Age-related clonal mutations and low-shedding tumors require careful assay design and expert interpretation.

Questions biomarkers can help answer

When you look at your pathology and lab reports, think about the questions they are designed to tackle:

• What is it? IHC panels, flow cytometry, and genetic testing identify the lymphoma subtype with precision.¹
• How fast is it? Proliferation indices, LDH, and clinical tempo set expectations for pace.²
• What drives it? Translocations (MYC, BCL2, BCL6), mutations (EZH2, MYD88, TP53), and viral markers (EBV) point to biology.
• Where is it? PET-CT or CT define distribution and extranodal involvement.
• How much is there? Stage, IPI-like scores, and serum markers outline burden.²
• Is it changing? PET response, ctDNA dynamics, MRD in CLL, and symptom shifts help flag remission, resistance, or transformation.³

A closer look at a few high-yield markers

CD20, CD19, CD3, and the basics: CD20 tags mature B cells, CD19 is present earlier in B-cell development and persists across most mature B-cell lymphomas, and CD3 marks T cells. This trio anchors lineage calls. Add CD10 and BCL6 to lean toward germinal center B cells, MUM1 to suggest late B-cell activation, and cyclin D1/SOX11 to support mantle cell lineage. These are the practical "GPS pins" for classification.

MYC, BCL2, BCL6 rearrangements: Detectable by FISH, these structural changes shift risk.⁶ ¹⁷ A single MYC translocation defines Burkitt when the morphology and phenotype fit. In DLBCL, pairing MYC with BCL2 and/or BCL6 rearrangements signals a highly proliferative biology with distinct outcomes in historical series. Protein overexpression by IHC picks up a broader group but does not always match the genetics, which is why both can appear in a report.

MYD88 L265P and EZH2 Y641: MYD88 L265P is a signature mutation in Waldenström macroglobulinemia and appears in a subset of ABC-type DLBCL.¹⁹ EZH2 mutations cluster in germinal center-driven diseases such as follicular lymphoma and some GCB DLBCL. Each adds diagnostic and mechanistic color and, in certain contexts, therapeutic relevance.

TP53 aberrations: Deletions and mutations affecting TP53 carry consistent adverse prognosis across NHL subtypes.¹⁴ When TP53 shows up, it is a cue that the tumor may behave independently of classic risk models.

How biomarkers fit into real care

Biomarkers inform rather than dictate. A simple way to picture it: your lymphoma "story" emerges from the intersection of pathology, genetics, blood tests, imaging, and how you feel. Two patients with the same translocation can have different courses if one has limited-stage disease and the other has widespread involvement. That is why doctors read biomarkers alongside the clinical narrative and imaging rather than as stand-alone verdicts.

When results conflict, the safest move is to reconcile them with additional data. If ctDNA suggests rising tumor DNA but PET-CT is quiet and you feel well, repeating tests, looking for hidden sites, or re-biopsy of a suspicious area may clarify. That measured pause is normal. In research settings, integrated models that combine imaging, ctDNA, and clinical scores are improving prediction — more data, smarter fusion.

What to expect in a high-quality pathology report

A strong report typically includes the diagnosis with the recognized entity name, a narrative of the morphology, a table or list of IHC stains with interpretation, flow cytometry results if performed, cytogenetic or FISH findings, and any molecular mutations assessed. It may also comment on proliferation (Ki-67), EBV status, and special features such as necrosis or sclerosis that help subclassify. When something is uncertain, the best reports state it plainly and recommend the next diagnostic step.

Where the field is heading

Three trends are worth watching:

• Deeper molecular profiling: Panel sequencing is moving from select cases to broader use, improving subtyping and opening trials.
• Liquid monitoring: ctDNA and blood-based MRD are inching closer to routine for certain subtypes, especially aggressive B-cell lymphomas and CLL.³
• Integrated risk models: Tools that merge imaging metrics, genomics, and clinical features are being tested to guide intensity of follow-up.

As with any fast-moving space, early signals need confirming studies. The pattern so far is consistent with other areas of oncology: better measurement yields better decisions.

Bottom line

Biomarkers turn a complex diagnosis into a map you can navigate. In NHL, they identify the exact subtype,¹ estimate risk,² and give objective ways to track response.³ No single test replaces the full picture, and there are real limits and caveats, from small biopsies to assay noise. When the data are read together by an experienced team, the insights are powerful. If you ever feel lost in the alphabet soup of CD markers and rearrangements, think of them as coordinates that tell your care team where they are and where they are going. The science is moving quickly, responsibly, and with a clear goal: more precise care, fewer surprises.