CLDN 18.2

IHC interpretation of CLDN18.2 expression

Accurate CLDN18.2 assessments are critical to guide treatment decision-making.1-4

Two researchers reviewing their findings
Two researchers reviewing their findings
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CLDN18.2-positivity is defined as ≥75% of tumor cells demonstrating moderate-to-strong (2+/3+) membranous CLDN18 staining, based on IHC assessments in two recently published global Phase 3 studies.1, 2

In G/GEJ tumors, assessment for CLDN18.2 expression is determined by the percentage of tumor cells with moderate-to-strong membrane staining3

  • Tissue slides with tumor cells present can demonstrate varying levels of CLDN18 membranous staining intensity, ranging from no staining to strong staining (0 to 3+)
Example CLDN18 Staining Intensities in G/GEJ cancer
Example CLDN18 Staining Intensities in G/GEJ cancer
Example CLDN18 Staining Intensities in G/GEJ cancer
Example CLDN18 Staining Intensities in G/GEJ cancer

Practical guidance on CLDN18.2 expression interpretation5*

Evaluation guidelines

Staining definition and intensity scoring

  • A cell is defined as CLDN18.2 positive in the presence of moderate-to-strong membranous staining (apical, circumferential, basolateral/lateral, or microluminal)
  • Staining of necrotic tissue/debris and staining of cytoplasm are considered negative; do not include in the score
  • Account for edge effect during interpretation
  • Staining intensity is scored between 0 and 3+ (absent, 0; weak, 1+; moderate, 2+; and strong, 3+)
  • A strong intensity is defined as dark brown to black immunoreactivity at low magnification (x4 objective) when 3,3'-diaminobenzidine is used as a chromogen

Heterogeneity and special scenarios

  • CLDN18.2 staining shows a high degree of intratumoral heterogeneity
    • CLDN18.2 expression may vary across different areas of the specimen; therefore, the report should represent an assessment of all components of the specimen
  • A low-magnification inspection of the stained slide should be completed to determine an overall impression of the staining pattern and heterogeneity of expression
  • In poorly cohesive carcinomas, assessment of both the CLDN18 stained slide and the matched hematoxylin and eosin-stained slide should be performed

Controls, inclusion, cutoff, and review

  • Normal gastric glands are CLDN18.2 positive and demonstrate strong membrane staining
  • Results should be considered invalid if a control sample demonstrates an absence of strong gastric epithelial staining, an absence of weak-to-moderate gastric epithelial staining in areas of metaplasia, or excessive nonspecific background staining
  • When evaluating infiltrating tumor cells, ensure other cell types (such as fibrotic cells or inflammatory cells) are not evaluated
  • Intestinal metaplasia may express CLDN18.2 but should not be included in scoring
  • Normal gastric epithelium and intestinal metaplasia may be used as appropriate controls in routine practice
  • Moderate-to-strong (2+/3+) positive membrane staining in ≥75% of tumor cells is the cutoff required for a sample to be considered CLDN18.2-positive
  • If any step of staining evaluation is not acceptable, repeat staining or request a new specimen

Staining considerations5*

Signet ring cells and dysplastic lesions

Signet ring cells with high expression, signet ring cells with nonspecific staining and dysplastic lesions showing CLDN18.2 expression
Signet ring cells with high expression, signet ring cells with nonspecific staining and dysplastic lesions showing CLDN18.2 expression
  • Exclude tumor cells with nonspecific staining that is obtrusive to interpretation of specific staining (including in signet ring cells, dysplastic lesions, etc)
  • Signet ring cells may be misinterpreted because of nonspecific staining in the cytoplasm
  • The concordance between CLDN18.2 expression of dysplastic lesions and their matched invasive components is unknown

Cytoplasmic vs nuclear staining

Membrane expression obscured by strong cytoplasmic staining and example of higher nuclear staining
Membrane expression obscured by strong cytoplasmic staining and example of higher nuclear staining
  • Exclude tumor cells with only cytoplasmic staining
  • Excessive cytoplasmic staining can obscure membrane staining

Aberrant positivity

Negative staining in inflammatory cells (true negative) and aberrant positivity in acellular mucus of mucinous adenocarcinoma (false positive)
Negative staining in inflammatory cells (true negative) and aberrant positivity in acellular mucus of mucinous adenocarcinoma (false positive)
  • Ignore aberrant positivity in inflammatory cells, nonneoplastic cells, or acellular mucus of mucinous adenocarcinoma

Preanalytical artifacts

Thermal artifacts
Thermal artifacts
  • • Artifacts: Histologic artifacts originating from sample processing and microtomy can complicate the determination of CLDN18 IHC status
    • • Artifacts may include underfixation/fixation gradients, edge effects, 3,3'-diaminobenzidine trapping, coverslip errors, lack of staining, light spots, tearing, folding, thermal artifacts, and more
  • • Antigen stability may impact staining interpretation, and could result from environmental or processing issues, such as improper storage or underfixation

Impact of necrosis on CLDN18.2 interpretation

Matched H&E and IHC stain of gastric
tumor necrosis
(10X magnification)

  • Nonspecific CLDN18 staining may be observed in necrosis, which can make interpretation challenging
  • Tumor cell staining in areas of necrosis or within intraluminal necrotic debris should be excluded from scoring

To evaluate CLDN18.2 expression, a minimum of 50 viable cells is required.5

*Some CLDN18.2 interpretation guidance and staining considerations include information developed from expert opinion.

Expert Case Review

Watch expert-guided case studies of CLDN18-stained slides

Gastric biopsy with benign gastric mucosa expert case review
Video

Gastric biopsy with benign gastric mucosa

Resection specimen with diffuse infiltrate expert case review
Video

Resection specimen with diffuse infiltrate

Gastric biopsy with poorly differentiated diffuse type adenocarcinoma expert case review
Video

Gastric biopsy with poorly differentiated diffuse type adenocarcinoma

Click to view stain gallery or test your knowledge via the stain quiz

CLDN18, claudin 18; CLDN18.2, claudin 18 isoform 2; H&E, hematoxylin and eosin; IHC, immunohistochemistry.

References:

  • 1. Shitara K, Xu RH, Ajani JA, et al. Global prevalence of claudin 18 isoform 2 in tumours of patients with locally advanced unresectable or metastatic gastric or gastroesophageal junction adenocarcinoma. Gastric Cancer. 2024;27(5):1058-1068. 
  • 2. Shah MA, Shitara K, Ajani JA, et al. Nat Med. 2023;29(8):2133-2141.
  • 3. Pellino A, Brignola S, Riello E, et al. Association of CLDN18 protein expression with clinicopathological features and prognosis in advanced gastric and gastroesophageal junction adenocarcinomas [published online October 26, 2021]. J Pers Med. 2021. Accessed April 3, 2026. https://www.mdpi.com/2075-4426/11/11/1095
  • 4. Rohde C, Yamaguchi R, Mukhina S, et al. Comparison of claudin 18.2 expression in primary tumours and lymph node metastases in Japanese patients with gastric adenocarcinoma. Jpn J Clin Oncol. 2019;49(9):870-876.
  • 5. VENTANA CLDN18 (43-14A) RxDx assay interpretation guide for gastric adenocarcinoma including gastroesophageal junction (GEJ). Tucson, AZ; 2024.