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CLDN2 / Claudin-2 · IHC design guide
Design Immunohistochemistry for CLDN2
Plan chromogenic CLDN2 IHC on paraffin sections with the IHC-validated antibody at 0.5–1 μg/mL (datasheet: RP1042 IHC-P). Assess membranous staining in gallbladder glands and kidney proximal tubules while accounting for presumed off-target staining (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
CLDN2 Immunohistochemistry Experimental Design Guide
Expected localisation, validated protocols, controls and antibodies — the at-a-glance facts below, then the full design guide.
Must know before staining
| Expected localisation | Membranous staining in gallbladder, kidney and GI (HPA tissue IHC) | |
| Staining pattern | Membranous gallbladder glands and kidney proximal tubules (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Gallbladder+1 more · see all | |
| Negative control | Adipose tissue+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent across sections (standard IHC practice; not target-specific) | |
| Caveat | Presumed off-target staining was observed and disregarded (HPA tissue IHC) | |
| Regulation | Expression regulation not established (UniProt) | |
| Isoform / epitope | No annotated isoforms; epitope side matters across 4 membrane spans (UniProt) |
Section 1
Recommended CLDN2 IHC & IF Protocols
The catalog antibody protocol is accompanied by published CLDN2 IHC protocols for goat mid jejunum (PMC4847856) and human lung tissue microarrays (PMC10366158).
| Sample | Tissue sections; selected-image fixative not specified (standard IHC workflow) |
| Fixation | Image fixative and duration unreported (datasheet RP1042); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat-induced epitope retrieval in citrate buffer, pH 6.0, 20 min at 95–98 °C (standard rule: cytoplasmic / membrane antigen) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% normal serum of the secondary host, 30 min, room temperature (standard) |
| Primary antibody | Rabbit anti-CLDN2, 0.5-1μg/ml (datasheet RP1042) |
| Primary incubation | Overnight at 4 °C (standard) |
| Detection | HRP-polymer secondary, DAB chromogen 5–10 min (standard) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | CLDN2-positive staining in glandular cells of gallbladder (HPA tissue IHC: High). HPA tissue profile: Membranous expression in gall bladder, kidney and GI. No signal in the no-primary control. |
💡Decision noteStart with citrate pH 6.0 HIER at 95–98 °C for 20 min (page antigen retrieval). The papers report EDTA pretreatment (PMC4847856) and autoclave retrieval (PMC10366158).
Section 2
What Is the Expected CLDN2 Staining Pattern?
CLDN2 is a 4-pass membrane protein at tight junctions (UniProt P57739 topology and subcellular location). In paraffin-section IHC, expect membranous staining in gallbladder glandular cells (High) and kidney proximal tubules (Medium); some GI glandular cells show lower staining (HPA tissue IHC). HPA rates the tissue profile Enhanced but reports medium staining–RNA consistency and presumed off-target binding that was disregarded (HPA tissue IHC).
| Gallbladder glandular cells show strong membrane-associated chromogen; kidney proximal tubule cells show a weaker membrane-associated pattern (HPA tissue IHC). | This fits the reported High gallbladder and Medium proximal-tubule staining (HPA tissue IHC). Junctional enrichment is biologically plausible because CLDN2 is a tight-junction membrane protein (UniProt P57739 subcellular location). |
| Staining is predominantly diffuse cytoplasmic or nuclear, with little membrane-associated signal (HPA tissue IHC expected profile). | Question that IHC pattern: HPA describes membranous tissue expression, and UniProt places CLDN2 at tight junctions and the cell membrane (HPA tissue IHC; UniProt P57739 subcellular location). HPA also reports additional nuclear locations in ICC-IF, so a nuclear observation warrants validation rather than automatic dismissal (HPA subcellular ICC-IF). |
| Strong signal appears in adipocytes or appendix glandular cells, or outside the expected cells in a positive section (HPA tissue IHC). | Those two cell populations are reported as Not detected (HPA tissue IHC). Check for cross-reactivity or endogenous detection activity as general IHC possibilities (general IHC practice); HPA also notes presumed off-target binding in its tissue assessment (HPA tissue IHC). |
| Chromogen spreads across tissue and blank areas, obscuring cell borders (general IHC practice). | Treat the distribution as background until controls clarify it (general IHC practice). A broad haze cannot establish the membranous glandular or proximal-tubule pattern reported for CLDN2 (HPA tissue IHC). |
| A gallbladder section lacks convincing glandular-cell staining (HPA tissue IHC expected profile). | Because gallbladder glandular cells are reported High, review the assay and tissue before calling the sample negative (HPA tissue IHC; general IHC practice). HPA’s Enhanced profile still has medium staining–RNA consistency and disregarded presumed off-target binding, so one section is not conclusive (HPA tissue IHC). |
💡Expected CLDN2 appearanceCall an IHC result consistent with CLDN2 when membrane-associated signal appears in gallbladder glandular cells at High intensity or kidney proximal tubule cells at Medium intensity; diffuse staining or strong signal in HPA Not detected cells warrants investigation (HPA tissue IHC; general IHC practice).
| Membrane topology and epitope location (UniProt P57739 topology) | CLDN2 has 4 transmembrane segments, extracellular loops and cytoplasmic regions (UniProt P57739 topology). Interpret staining against its membrane and junction location; the supplied sources do not identify the catalog antibody’s epitope or establish a CLDN2-specific retrieval condition (UniProt P57739 subcellular location; supplied evidence). |
| Tissue and cell selection (HPA tissue IHC) | Gallbladder glandular cells provide a High reference; kidney proximal tubules are Medium (HPA tissue IHC). Stomach, duodenum, small-intestine and seminal-vesicle glandular cells are Low, so a faint result there is less decisive (HPA tissue IHC). |
| IHC evidence and antibody choice (HPA antibodies; HPA tissue IHC) | The IHC-validated antibody CAB002609 is rated Enhanced for IHC (HPA antibodies). HPA defines Enhanced through independent-antibody agreement or orthogonal data, while its tissue profile still notes medium staining–RNA consistency and disregarded presumed off-target binding (HPA antibody validation; HPA tissue IHC). |
| IF/ICC Q&A: What localisation should IF show? (HPA subcellular ICC-IF) | Mainly plasma membrane and cell junctions, both Supported; nucleoplasm, nucleoli and vesicles are additional Approved locations (HPA subcellular ICC-IF). The listed ICC ratings differ: HPA051548 is Supported and HPA074528 is Uncertain (HPA antibodies). Use the separate IF/ICC guide for its workflow (application scope). |
| Situation | Likely cause | Next action |
|---|---|---|
| Gallbladder glandular cells are blank or very faint despite their reported High level (HPA tissue IHC). | A failed staining run, unsuitable detection or tissue quality can remove an expected signal (general IHC practice). | Check a known-positive control, reagent performance and tissue morphology; review the validated IHC procedure’s retrieval and detection steps before interpreting absence (general IHC practice; HPA tissue IHC). |
| The section stains broadly, including blank areas (general IHC practice). | Nonspecific reagent binding or detection background may obscure tissue localisation (general IHC practice). | Inspect a negative reagent control, blocking, washing and chromogen development; assess specific signal only after membrane-associated cell borders are readable (general IHC practice; HPA tissue IHC). |
| Strong chromogen appears in adipocytes or appendix glandular cells, both reported Not detected (HPA tissue IHC). | Cross-reactivity or endogenous detection activity is possible; HPA reports presumed off-target binding in its tissue assessment (general IHC practice; HPA tissue IHC). | Compare a negative control and an expected-positive tissue, then reassess whether staining follows cell membranes and the reported cell distribution (general IHC practice; HPA tissue IHC). |
| Only nuclear or diffuse cytoplasmic staining is visible in paraffin-section IHC (HPA tissue IHC expected profile). | The compartment conflicts with HPA’s membranous tissue profile; ICC-IF additionally lists nuclear locations, so the discrepancy needs investigation (HPA tissue IHC; HPA subcellular ICC-IF). | Review morphology, controls and antibody validation before assigning it to CLDN2; seek membrane-associated staining in expected cells (general IHC practice; HPA tissue IHC). |
| Stomach or small-intestine glandular cells stain weakly (HPA tissue IHC). | Both are reported Low, making a faint signal compatible with the reference profile (HPA tissue IHC). | Judge localisation and controls, and compare with High gallbladder glandular cells rather than increasing development solely to make a Low tissue look strong (HPA tissue IHC; general IHC practice). |
| Kidney staining is weaker than gallbladder staining (HPA tissue IHC). | The reported levels differ: proximal tubules are Medium and gallbladder glandular cells are High (HPA tissue IHC). | Check that kidney signal is in proximal tubule cells and associated with membranes; investigate only if localisation or control performance fails (HPA tissue IHC; general IHC practice). |
Experiment evidence
IHC & IF images showing CLDN2 staining
RP1042 · IHCAnti-Claudin 2 antibody, RP1042, IHC(P) IHC(P): Human Intestinal Cancer Tissue
A03033-1 · IFIF analysis of Claudin 2/CLDN2 using anti-Claudin 2/CLDN2 antibody (A03033-1). Claudin 2/CLDN2 was detected in an immunocytochemical section
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000165376-CLDN2/tissue/cerebral+cortex#imid_810992
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000165376-CLDN2/tissue/colon#imid_810910
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000165376-CLDN2/tissue/liver#imid_810919
HPA · GallbladderHuman Protein Atlas IHC, Gallbladder. Source: https://www.proteinatlas.org/ENSG00000165376-CLDN2/tissue/gallbladder#imid_810887
HPA · ICC-IF A-549Human Protein Atlas immunofluorescence, A-549, antibody HPA051548; green = CLDN2, blue = nucleus, red = microtubules.
HPA · ICC-IF A-549Human Protein Atlas immunofluorescence, A-549, antibody HPA051548; green = CLDN2, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for CLDN2 IHC & IF
🧪Run gallbladder first: its glandular cells should show strong CLDN2 staining (HPA: High in gallbladder glandular cells). Use appendix glandular cells as a negative comparator (HPA: Not detected in appendix glandular cells); on the gallbladder slide, assess nonglandular areas for background rather than assuming their CLDN2 status from the glandular-cell result (HPA: gallbladder glandular-cell result only).
Positive control tissue: Gallbladder (Glandular cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show CLDN2 in A-549, CACO-2, U2OS, RPTEC/TERT1, hTERT-RPE1 (serum starved), with annotated localisation: Plasma membrane (supported), Cell Junctions (supported) (HPA subcellular).
Technical controls: Include no-primary (secondary-only), host-species- and isotype-matched antibody, and CLDN2-knockout biological controls (standard IHC practice). For chromogenic gallbladder IHC, block endogenous peroxidase and check for pigment that could be mistaken for signal; block endogenous biotin if using avidin–biotin detection (standard IHC practice).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected RP1042 paraffin-IHC caption does not state a fixative (caption: IHC(P), fixative not stated). Retrieval dependence is unreported, so optimize antigen retrieval empirically for the paraffin sections (standard IHC practice). The supplied evidence does not establish that frozen sections or IF are easier; for gallbladder, distinguish chromogen from tissue pigment using the control slide (caption: IHC(P); standard IHC practice).
HPA tissue IHC evidence for CLDN2
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Enhanced — Medium consistency between antibody staining and RNA expression data. Presumed off target binding observed and disregarded.). Rows are taken directly from the HPA tissue chart — click any row's HPA link to view the source.
Positive expression · recommended positive controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Gallbladder | Glandular cells | High | Protein (IHC) | HPA → |
| Kidney | Proximal tubules (cell body) | Medium | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Appendix | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | Not detected | Protein (IHC) | HPA → |
| Caudate | Glial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced CLDN2 IHC Tips
Troubleshoot CLDN2 chromogenic IHC by checking retrieval, tissue preparation, junctional staining and cell type before interpreting signal strength.
What retrieval should I try first if CLDN2 staining is weak?
Start with citrate pH 6.0 heat retrieval at 95–98 °C for 20 min (page IHC retrieval setting). CLDN2 spans the membrane 4 times, so assess whether staining outlines cell junctions rather than judging intensity alone (UniProt P57739 topology; UniProt P57739 localisation). If signal remains weak, test EDTA pH 9 on an adjacent section as a fallback, keeping antibody concentration and detection constant (standard IHC optimization). Compare both conditions with a positive control such as gallbladder glandular cells, which show high staining, and a no-primary control (HPA tissue IHC; standard IHC practice). Record tissue damage or diffuse staining alongside signal gain (standard IHC practice).
Could fixation explain weak or uneven CLDN2 staining?
Target-specific fixation sensitivity is unknown: the RP1042 IHC(P) caption identifies human intestinal cancer tissue but does not state its fixative (RP1042 caption). For paraffin sections, record the fixative and fixation duration, then compare sections prepared under documented conditions before changing antibody concentration (standard IHC practice). Excessive or uneven fixation can alter antigen accessibility in IHC generally; this is a troubleshooting possibility, not an established CLDN2-specific effect (standard IHC practice). Keep citrate pH 6.0 retrieval at 95–98 °C for 20 min constant during that comparison (page IHC retrieval setting). Judge recovery by cellular pattern and controls, not intensity alone (standard IHC practice).
Where should a convincing CLDN2 chromogenic signal appear?
Expect staining at epithelial cell membranes and junctions: CLDN2 is a tight-junction membrane protein, and HPA reports membranous expression in gallbladder, kidney and gastrointestinal tissue (UniProt P57739 localisation; HPA tissue IHC). Check whether signal follows boundaries between appropriate cells rather than pooling over whole sections (standard IHC interpretation). HPA also reports additional nucleoplasmic, nucleolar and vesicular localisation in cell imaging, so an isolated intracellular IHC pattern needs independent validation before assignment to CLDN2 (HPA subcellular; standard IHC interpretation). Use gallbladder glandular cells as a high-staining comparison and kidney proximal tubules as a medium-staining comparison (HPA tissue IHC). Document the compartment and cell type separately from intensity (standard IHC practice).
How should unknown epitope position affect CLDN2 IHC troubleshooting?
UniProt annotates one CLDN2 chain of 230 residues and 0 isoforms; that annotation does not establish where the catalog antibody binds (UniProt P57739 processing; UniProt P57739 isoforms). The protein has 2 extracellular loops and a cytoplasmic C terminus spanning residues 184–230 (UniProt P57739 topology). Phosphoserines are annotated at residues 219 and 223, but their effect on this antibody's staining is unknown (UniProt P57739 modified residues). Check the antibody's stated immunogen or epitope before attributing failed retrieval to a particular loop or terminal region (standard IHC validation practice). If the epitope remains unspecified, report staining as antibody dependent (standard IHC reporting practice).
How can I check a CLDN2 pattern by IF alongside chromogenic IHC?
For a separate IF experiment, multiplex CLDN2 with an independently validated marker of the expected glandular or proximal-tubule cell population, then inspect junctional overlap within those cells (HPA tissue IHC; UniProt P57739 localisation; standard IF practice). Choose a far-red fluorophore when tissue autofluorescence interferes with shorter-wavelength channels, and include unstained and single-label controls (standard IF practice). CLDN2 has extracellular loops and cytoplasmic regions, so set permeabilisation according to the antibody's documented epitope side rather than assuming access through an intact membrane (UniProt P57739 topology; standard IF practice). If that side is unknown, compare permeabilised and nonpermeabilised preparations while holding fixation constant (standard IF optimization). IF results require their own antibody validation (standard IF practice).
What should I check when CLDN2 IHC looks diffusely brown?
First compare the section with a no-primary control and check whether color appears outside cell boundaries or at tissue edges (standard chromogenic IHC practice). Quench endogenous peroxidase before DAB detection and use an appropriate protein block, recording the conditions used for each run (standard chromogenic IHC practice). If background persists, titrate the IHC-validated antibody and detection reagents separately while keeping citrate pH 6.0 retrieval fixed (standard IHC optimization; page IHC retrieval setting). HPA reports presumed off-target binding that was disregarded in its tissue assessment, so diffuse signal deserves careful control-based review (HPA tissue IHC reliability description). Score junctional staining separately from diffuse background (UniProt P57739 localisation; standard IHC practice).
How should I quantify CLDN2 across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the eligible cell population and junctional compartment before scoring, since CLDN2 is a tight-junction membrane protein (UniProt P57739 localisation; standard IHC quantification practice). Report the percentage of positive eligible cells and a prespecified intensity score; an H-score combines percentage and intensity on a 0–300 scale (standard IHC scoring practice). If measuring stained junction length or area, normalize it to epithelial length or eligible epithelial area, respectively, rather than total section area (standard image-analysis practice). Exclude folds, necrosis and section edges using the same rules for all samples (standard IHC quantification practice). Record control staining and acquisition settings alongside each score (standard IHC quantification practice).
How do I distinguish true CLDN2 signal from artefact?
Favor a membrane or junctional pattern in the expected epithelial cells, consistent with CLDN2 localisation and HPA's tissue profile (UniProt P57739 localisation; HPA tissue IHC). Gallbladder glandular cells show high staining and kidney proximal tubules medium staining, while appendix glandular cells were not detected in the HPA assessment (HPA tissue IHC). Treat staining confined to cut edges, necrotic areas or the no-primary control as possible processing or detection artefact (standard IHC interpretation). Investigate isolated nuclear or diffuse cytoplasmic staining before calling a sample positive, although HPA cell imaging lists additional intracellular locations (HPA subcellular; standard IHC interpretation). Confirm uncertain patterns with a second validated assay or antibody (standard IHC validation practice).
Boster reagents
Best CLDN2 / Claudin-2 IHC Antibodies
Two anti-CLDN2 antibodies have real application images: paraffin-section IHC in human intestinal cancer tissue (RP1042 image caption) and IF/ICC in human Caco-2 cells (A03033-1 image caption).
Real IHC data
Anti-Claudin 2/CLDN2 Antibody ®
Cat # RP1042
Real IF data
Anti-Claudin 2/CLDN2 Antibody ®
Cat # A03033-1
RP1042 will render with human intestinal cancer tissue IHC data from a paraffin section (RP1042 image caption). A03033-1 will render with IF/ICC data from human Caco-2 cells (A03033-1 image caption).
Which to pick: Choose RP1042 for paraffin-section tissue IHC (RP1042 applications and image caption); its fixative is unreported (RP1042 image caption). Choose A03033-1 for human IF/ICC (A03033-1 applications, reactivity and image caption). For rat samples, RP1042 lists rat reactivity, but its IHC image and dilution data concern human samples; clonality is unreported for both antibodies (catalog payload).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P57739 (CLD2_HUMAN, Claudin-2).
- Human Protein Atlas. CLDN2 tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. CLDN2 subcellular location (ICC-IF): Mainly localized to the plasma membrane and cell junctions. In addition localized to the nucleoplasm, nucleoli and vesicles..
- Human Protein Atlas. CLDN2 antibody validation summary (3 antibodies).
- Expression of Tight Junction Proteins and Cadherin 17 in the Small Intestine of Young Goats Offered a Reduced N and/or Ca Diet. PloS one 2016 — PMC4847856.
- Multidirectional characterization of cellular composition and spatial architecture in human multiple primary lung cancers. Cell death & disease 2023 — PMC10366158.
- Claudin-2 protects against colitis-associated cancer by promoting colitis-associated mucosal healing. The Journal of clinical investigation 2023 — PMC10688979.
- Inflammation-Induced Claudin-2 Up-Regulation Limits Pancreatitis Development by Enhancing Pancreatic Ductal Transport. Gastroenterology 2026 — PMC13222116.
- PubMed PMID:11934881 — UniProt-cited evidence.
- PubMed PMID:12975309 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for CLDN2 (Claudin-2). Expected localisation Membranous staining in gallbladder, kidney and GI (HPA tissue IHC). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: RP1042 at 0.5-1μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Gallbladder (Glandular cells); negative Adipose tissue. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Tissue sections; selected-image fixative not specified (standard IHC workflow); 2. Fixation: Image fixative and duration unreported (datasheet RP1042); verify before use.; 3. Sectioning: 4–5 µm sections on charged slides (standard); 4. Deparaffinisation: Xylene, graded ethanol series to water (standard); 5. Antigen retrieval: Heat-induced epitope retrieval in citrate buffer, pH 6.0 , 20 min at 95–98 °C…; 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% normal serum of the secondary host, 30 min , room temperature (standard); 8. Primary antibody: Rabbit anti-CLDN2, 0.5-1μg/ml (datasheet RP1042); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across sections (standard IHC practice; not target-specific); Caveat: Presumed off-target staining was observed and disregarded (HPA tissue IHC); Isoform / epitope: No annotated isoforms; epitope side matters across 4 membrane spans (UniProt); Regulation: Expression regulation not established (UniProt). Evidence: UniProt P57739, HPA tissue IHC · Enhanced, HPA ICC-IF · Mainly localized to the plasma membrane and cell junctions. In…, Datasheet RP1042, PMC4847856, PMC10366158. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/CLDN2/immunohistochemistry-ihc-guide.