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PA19.1-6 | Hodgkin Lymphoma — Gross & Microscopic Identification — Part 3
Mixed Cellularity — Detailed Histology
Mixed Cellularity Classical Hodgkin Lymphoma: Detailed Histology
Mixed cellularity (MC) is the second most common HL subtype (20–25%) and the EBV-associated subtype most commonly tested in tropical/resource-limited settings.
Histological hallmarks:
• Diffuse or vaguely nodular effacement of nodal architecture — no broad collagen bands
• Classic binucleate RS cells are numerous and easy to find (more RS cells per field than NS)
• Polymorphous background with eosinophils, plasma cells, histiocytes, and lymphocytes in roughly equal proportions
• Mild interstitial fibrosis (fine reticulin), but NO broad collagen septa
• EBV-encoded RNA (EBER) detectable by ISH in 50–75% of cases
Key distinction from NS:
| Feature | NS | MC |
|---|---|---|
| Collagen bands | Broad, birefringent | Absent |
| Dominant cell | Lacunar cell | Classic RS cell |
| Background | Mixed, less eosinophil-heavy | Eosinophil/plasma-cell-rich |
| EBV association | ~10–25% | ~50–75% |
| Age/site | Young adults, mediastinum | Older adults/paediatric, peripheral nodes |
Clinical note: MC is the predominant subtype in HIV-positive patients and in sub-Saharan Africa and South/Southeast Asia.
Immunohistochemistry Panel for HL
IHC Confirmation Panel for Classic Hodgkin Lymphoma
When morphology alone is insufficient — or in any examination question about confirmation — apply the standard IHC panel:
Classic Hodgkin Lymphoma (cHL) profile:
| Marker | Result | Interpretation |
|---|---|---|
| CD30 | Positive (membranous + Golgi) | Activation marker; most sensitive for cHL neoplastic cells |
| CD15 | Positive | Granulocyte antigen aberrantly expressed on RS cells; high specificity |
| CD45 (LCA) | Negative | Loss of common leukocyte antigen distinguishes RS cells from reactive lymphocytes |
| PAX5 | Positive (dim) | Dim nuclear staining; confirms B-cell lineage despite CD20 loss |
| CD20 | Negative (rare weak positivity in ~10%) | Useful to exclude DLBCL |
| CD3 | Negative | Excludes T-cell lymphoma |
NLPHL profile (contrast):
| Marker | Result |
|---|---|
| CD20 | Positive (strong) |
| CD45 | Positive |
| CD30 | Negative |
| CD15 | Negative |
| OCT2, BOB1 | Positive (retained B-cell transcription factors) |
The most tested IHC question: 'What IHC profile differentiates cHL from DLBCL?' Answer: cHL is CD30+, CD15+, CD45−; DLBCL is CD20+, CD45+, CD30− (usually).
Practical note: IHC is performed on the same paraffin block. You will not be asked to run the stains in the practical, but you must be able to interpret a panel result and explain what each marker confirms or excludes.
SELF-CHECK
IHC on a lymph node biopsy shows: CD30+, CD15+, CD45−, CD20−, PAX5 dim+. Which diagnosis does this profile confirm?
A. Classic Hodgkin lymphoma
B. Diffuse large B-cell lymphoma
C. Nodular lymphocyte-predominant Hodgkin lymphoma
D. Anaplastic large cell lymphoma
Reveal Answer
Answer: A. Classic Hodgkin lymphoma
CD30+/CD15+/CD45−/CD20−/PAX5-dim is the canonical IHC fingerprint of classic Hodgkin lymphoma. DLBCL is CD20+/CD45+/CD30−. NLPHL is CD20+/CD45+/CD30−/CD15−. ALCL is CD30+ but ALK+ and CD15−. The CD45 negativity is key — it separates the RS cell from virtually all reactive lymphoid cells.
H5P Practical Activity: 2×2 Composite Identification Grid
The following image contains four panels for the H5P Image Hotspots identification exercise. For each panel, identify the labelled structure and answer the embedded question.
Hodgkin Lymphoma Histological Features: Reed-Sternberg Cells and Tissue Architecture
Panel guide (use when reviewing after the H5P activity):
• Panel A — Classic RS cell: look for the two mirror-image eosinophilic nucleoli. CD30+/CD15+ on IHC.
• Panel B — Lacunar cell: the clear space (lacuna) is formalin-fixation artefact; nucleus is lobulated, not mirror-image.
• Panel C — NS bands: broad, pale, birefringent collagen. Compare to the thin reticulin fibrosis of MC.
• Panel D — MC background: the 'zoo' of cell types — eosinophils (bright pink granules), plasma cells (clockface nucleus), histiocytes (pale kidney-shaped nucleus), lymphocytes (small, dark). Find the RS cell in the crowd.
CLINICAL PEARL
The 1–5% rule and the searchability problem: In a mixed cellularity slide, RS cells constitute less than 5% of all cells. A student who stops scanning after finding one RS cell has done the minimum. The examiner wants you to demonstrate systematic scanning: raster pattern at 20×, confirm at 40×, note the background. In an MCQ, if asked 'what percentage of the tumour mass is made up of Reed-Sternberg cells?' the answer is 1–5% — the rest is reactive stroma. This is also why flow cytometry fails to detect HL (too few neoplastic cells to gate reliably) and why tissue biopsy is mandatory for diagnosis.
SELF-CHECK
Why is flow cytometry unreliable for diagnosing Hodgkin lymphoma, and what specimen type is mandatory?
A. RS cells constitute only 1–5% of the tumour; tissue biopsy (excisional) is mandatory for morphological identification
B. HL cells express no surface antigens detectable by flow cytometry; fine-needle aspiration is sufficient
C. Flow cytometry requires live cells; HL cells are always necrotic at diagnosis
D. HL is a T-cell tumour and flow cytometry panels only detect B-cell markers
Reveal Answer
Answer: A. RS cells constitute only 1–5% of the tumour; tissue biopsy (excisional) is mandatory for morphological identification
The neoplastic RS cells are so sparse (1–5% of the tumour) that they are below the detection threshold of flow cytometry, which requires sufficient numbers of phenotypically abnormal cells to gate. Excisional or core-needle biopsy preserving architecture is essential — FNA is unreliable because it disrupts the architecture needed to see the RS cell in its polymorphous reactive context, and it cannot demonstrate nodular architecture or collagen bands.