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PA6.4-6 | Tumour Effects, Immunology & Laboratory Diagnosis — SDL Guide (Part 3)

Immunohistochemistry (IHC)

Immunohistochemistry uses specific antibodies applied to formalin-fixed, paraffin-embedded (FFPE) tissue sections to detect antigens in situ. Chromogen (DAB — brown) or fluorescent tag visualises the result.

IHC solves critical diagnostic problems:

Tumour lineage (where did this come from?):

Marker	Cell/tumour type identified
Cytokeratin (CK)	Epithelial origin (carcinomas)
LCA (CD45)	Lymphoid origin (lymphomas)
S100 protein	Neural crest — melanoma, nerve sheath tumours
Desmin / myogenin	Muscle tumours (rhabdomyosarcoma)
Vimentin	Mesenchymal origin (sarcomas)
TTF-1	Lung or thyroid primary
PSA	Prostate carcinoma

Identifying cancer of unknown primary (CUP): IHC panel (CK7/CK20/TTF-1/CDX2/PSA) narrows the primary site when metastasis presents first.

3. Prognostic/predictive markers that guide therapy:
• ER/PR (oestrogen/progesterone receptor) in breast cancer → eligibility for hormonal therapy.
• HER2 (ERBB2) in breast/gastric cancer → eligibility for trastuzumab.
• Ki-67 — proliferation index; high Ki-67 = aggressive tumour.
• PD-L1 — predicts response to checkpoint inhibitors.

A three-panel medical infographic explains IHC workflow on FFPE tissue and maps common immunohistochemical markers to tumour lineage and likely primary sites. — **Panel A:** FFPE tissue block, tissue section on glass slide, tumour antigen, primary antibody, secondary antibody with enzyme or fluorescent tag, DAB brown positive staining, fluorescent signal, negative background cells. **Panel B:** Unknown tumour cell cluster, tumour lineage question, carcinoma, lymphoma, melanoma or nerve sheath tumour, muscle tumour, sarcoma. **Panel C:** Cytokeratin CK for epithelial origin or carcinoma, LCA CD45 for lymphoid origin or lymphoma, S100 protein for neural crest tumours including melanoma and nerve sheath tumour, desmin or myogenin for skeletal muscle differentiation and rhabdomyosarcoma, vimentin for mesenchymal origin or sarcoma, TTF-1 for lung or thyroid primary, PSA for prostate primary.

SELF-CHECK

A 55-year-old woman presents with a 2-cm lymph node mass. Biopsy shows large atypical cells. IHC reveals: CK-positive, LCA-negative, S100-negative. What is the most likely cell lineage?

A. Lymphoma (B-cell origin)

B. Melanoma (neural crest origin)

C. Carcinoma (epithelial origin)

D. Rhabdomyosarcoma (muscle origin)

Reveal Answer

Answer: C. Carcinoma (epithelial origin)

Cytokeratin (CK) positivity marks epithelial origin — i.e., a carcinoma (likely metastatic). LCA (CD45) positivity would indicate lymphoma; S100 positivity would suggest melanoma or nerve sheath tumour. This IHC profile is classic for metastatic carcinoma in a lymph node.

Tumour Markers

Tumour markers are molecules — proteins, hormones, or enzymes — produced by tumour cells or by the host in response to the tumour, detectable in blood (or other fluids).

Critical principle: Tumour markers are generally NOT reliable for screening (low specificity — raised in benign conditions) but are highly valuable for monitoring treatment response and detecting recurrence after therapy.

Infographic explaining tumour markers as tumour- or host-derived molecules mainly used for monitoring therapy response and recurrence, with a timeline graph and summary table of major markers. — **Panel A:** Tumour mass, blood vessel, tumour-derived molecules, host response molecules, proteins, hormones, enzymes, principle that markers are not reliable screening tests but useful for monitoring and recurrence.. **Panel B:** Baseline marker level, treatment, falling marker level indicating response, follow-up monitoring, rising marker level suggesting recurrence.. **Panel C:** AFP, CEA, PSA, CA-125, CA19-9, hCG, Calcitonin, LDH with marker type, key associated tumours, clinical use, and benign causes..

Marker	Key tumour(s)	Clinical use	Benign causes
AFP (Alpha-fetoprotein)	Hepatocellular carcinoma (HCC); testicular germ cell (non-seminoma)	Diagnosis + monitoring	Hepatitis, cirrhosis; pregnancy
CEA (Carcinoembryonic antigen)	Colorectal carcinoma; gastric, pancreatic	Monitoring recurrence post-surgery	Smoking, IBD, cirrhosis
PSA (Prostate-specific antigen)	Prostate carcinoma	Screening (controversial) + monitoring	BPH, prostatitis
CA-125	Ovarian carcinoma	Monitoring; second-line screening in high risk	Endometriosis, PID, liver disease
CA19-9	Pancreatic carcinoma; biliary	Monitoring	Pancreatitis, cholangitis
β-hCG	Gestational trophoblastic disease; germ cell tumours	Diagnosis + monitoring	Pregnancy
Calcitonin	Medullary thyroid carcinoma (MTC)	Diagnosis + screening in MEN2	—
LDH	Lymphoma, leukaemia, testicular	Disease burden / prognosis	Haemolysis, MI, myopathy

PSA nuance: PSA is organ-specific (prostate), not cancer-specific. Raised in BPH and prostatitis. Its value lies in monitoring post-prostatectomy or post-radiotherapy (undetectable PSA = remission; rising PSA = recurrence).

Molecular and Cytogenetic Diagnosis

A multi-panel medical education diagram summarizing how FISH, PCR, NGS, and flow cytometry detect tumour genetic or antigenic abnormalities to guide diagnosis, prognosis, and targeted therapy. — **Panel A:** Tumour biopsy sample, molecular testing workflow, confirm diagnosis, predict prognosis, targeted therapy, companion diagnostic.. **Panel B:** FISH probe, interphase nucleus, chromosome locus, HER2 amplification, BCR-ABL translocation, deletion signal.. **Panel C:** DNA/RNA template, PCR amplification, RT-PCR, BCR-ABL fusion transcript, quantitative monitoring in CML, EGFR mutation in NSCLC.. **Panel D:** Tumour DNA fragments, NGS gene panel, sequencing reads, mutation calls, tumour mutation burden, immunotherapy response prediction.. **Panel E:** Cells in suspension, laser beam, fluorescent antibodies, CD markers, B-cell population, T-cell population, NK-cell population, leukaemia/lymphoma immunophenotyping.. **Panel F:** HER2 amplification, breast cancer, gastric cancer, trastuzumab; BCR-ABL fusion, CML, tyrosine kinase inhibitor; EGFR mutation, NSCLC, EGFR inhibitor; high TMB, immunotherapy..

Molecular diagnostics identify specific genetic alterations that (a) confirm diagnosis, (b) predict prognosis, and (c) determine eligibility for targeted therapy (companion diagnostics).

Key techniques:

FISH (Fluorescence In Situ Hybridisation): Fluorescent DNA probes hybridise to specific chromosomal loci on tissue sections or smears. Detects gene amplification (HER2 amplification in breast cancer), translocations (BCR-ABL in CML), and deletions.

PCR (Polymerase Chain Reaction): Amplifies specific DNA/RNA sequences. RT-PCR detects BCR-ABL fusion transcript (quantitative monitoring in CML); EGFR mutation testing in NSCLC.

NGS (Next-Generation Sequencing): Simultaneous sequencing of hundreds of cancer-relevant genes. Enables tumour mutation burden (TMB) assessment — predicts response to immunotherapy.

Flow cytometry: Quantifies surface antigens on cells in suspension (blood, marrow, effusions). Critical for immunophenotyping leukaemias and lymphomas (B-cell vs T-cell vs NK, CD markers).

Key companion diagnostic examples:

Molecular finding	Tumour	Targeted therapy unlocked
HER2 amplification	Breast, gastric	Trastuzumab (Herceptin)
EGFR mutation	Non-small cell lung (adenocarcinoma)	Erlotinib, gefitinib
BCR-ABL (t(9;22))	CML	Imatinib (Gleevec)
BRAF V600E	Melanoma	Vemurafenib
PD-L1 expression	Multiple solid tumours	Pembrolizumab

Molecular testing has shifted oncology from 'site of origin' to 'molecular phenotype' — the same EGFR mutation in lung and colon may respond to the same drug.