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IRF2BP1 / Interferon regulatory factor 2-binding protein 1 · IHC design guide
Design Immunohistochemistry for IRF2BP1
Plan chromogenic IHC for IRF2BP1 around its general nuclear tissue staining (HPA tissue IHC) and a primary antibody range of 2–5 μg/ml (datasheet A11406-2). Adrenal glandular cells and bone marrow hematopoietic cells show high staining and can serve as reference populations (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
IRF2BP1 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 | General nuclear staining observed (HPA tissue IHC) | |
| Staining pattern | Nuclear staining across tissues (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A11406-2) | |
| Positive control | Adrenal gland+4 more · see all | |
| Negative control | Heart muscle |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent across sections. (standard IHC practice; not target-specific) | |
| Caveat | Staining has medium consistency with RNA expression (HPA tissue IHC) | |
| Regulation | No expression regulator annotated (UniProt) | |
| Isoform / epitope | One full-length chain; no isoforms annotated (UniProt) |
Section 1
Recommended IRF2BP1 IHC & IF Protocols
The catalog antibody’s IHC-P protocol (datasheet: A11406-2) is accompanied by one published colon carcinoma protocol (PMC4192020).
| Sample | Paraffin-embedded human acinic cell carcinoma of parotid cancer tissue; fixative not specified (datasheet A11406-2) |
| Fixation | Image fixative and duration unreported (datasheet A11406-2); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat retrieval: EDTA pH 8.0 (datasheet A11406-2); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A11406-2) |
| Primary antibody | Rabbit anti-IRF2BP1, 2-5 μg/ml (datasheet A11406-2) |
| Primary incubation | Overnight at 4 °C (datasheet A11406-2) |
| Detection | HRP-conjugated secondary, DAB chromogen (datasheet A11406-2) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | IRF2BP1-positive staining in glandular cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: General nuclear expression. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA retrieval at pH 8.0 for the catalog antibody (datasheet: A11406-2); the article does not specify its retrieval solution (PMC4192020).
Section 2
What Is the Expected IRF2BP1 Staining Pattern?
IRF2BP1 is a nuclear protein with no transmembrane segment (UniProt Q8IU81 topology and subcellular location). In paraffin-section IHC, expect nuclear staining across multiple cell types, including adrenal and intestinal glandular cells, cervical squamous epithelial cells, and bone-marrow hematopoietic cells (HPA tissue IHC). HPA describes general nuclear expression, but rates its tissue IHC profile Approved with medium consistency between staining and RNA data, pending external verification (HPA tissue IHC).
| Distinct nuclear chromogen in colon glandular cells, with nuclei identifiable on the counterstain (HPA: High in colon glandular cells). | This fits the expected compartment and an observed high-staining cell population (UniProt Q8IU81: nucleus; HPA: High in colon glandular cells). Score the relevant nuclei, noting the proportion and intensity separately; a high HPA category does not prescribe one intensity for every section. |
| Predominantly cytoplasmic or membranous staining, with little nuclear signal. | Treat this as discordant with IRF2BP1's nuclear location and lack of a transmembrane segment (UniProt Q8IU81 topology and subcellular location). Review morphology and detection controls before interpreting it as IRF2BP1; compartment mismatch alone does not identify the source of artefact. |
| Prominent chromogen in an unexpected cell population, while expected nuclei are weak or clear. | Check cell identity against the tissue profile before calling it target staining (HPA tissue IHC: general nuclear expression; High in specified cell populations). Cross-reactivity or endogenous detection activity are possible IHC explanations; use appropriate reagent and detection controls to distinguish them. |
| Diffuse color over nuclei, cytoplasm, and surrounding tissue, without crisp cellular boundaries. | This is difficult to score as nuclear IRF2BP1 (UniProt Q8IU81: nucleus; HPA tissue IHC: general nuclear expression). In routine IHC, uneven blocking, residual detection reagents, or excessive chromogen development can contribute to background; compare control sections and assess the detection run. |
| No nuclear signal in an adequately sampled adrenal gland or colon section (HPA: High in adrenal and colon glandular cells). | A technical failure is plausible, but HPA's High category is an observed profile, not a guarantee for every specimen (HPA tissue IHC; Approved, medium consistency, pending external verification). Check the IHC-validated antibody, detection controls, and tissue integrity before reporting absence. |
💡Expected IRF2BP1 appearanceCall a result consistent when identifiable nuclei stain in an HPA-reported positive cell population, potentially strongly in its High groups; predominantly cytoplasmic, membranous, or diffuse color is suspect (HPA tissue IHC: general nuclear expression and High groups; UniProt Q8IU81: nucleus).
| Compartment and topology | Use nuclear localization as the primary reading criterion: UniProt places IRF2BP1 in the nucleus and reports no transmembrane segment (UniProt Q8IU81). These annotations support interpretation of the pattern; they do not establish a retrieval condition. |
| Tissue and cell context | HPA reports low tissue RNA specificity and general nuclear IHC expression, with High staining in several listed cell populations, Low in tonsil germinal-center cells, and Not detected in heart cardiomyocytes (HPA tissue IHC). Interpret the named cell population within each tissue. |
| Strength of IHC evidence | The tissue profile is Approved, with medium staining-to-RNA consistency and external verification pending; HPA042164 is IHC Approved (HPA tissue IHC; HPA antibodies). These labels support a reference pattern but do not make an unexpected stain conclusive. |
| Processing and modifications | UniProt reports one chain spanning residues 1–584, no signal peptide, and several modified residues (UniProt Q8IU81). These facts do not identify the antibody epitope or predict how paraffin processing, fixation, or antigen retrieval affects detection. |
| IF/ICC Q: What pattern should a separate IF/ICC assay seek? | A: Nucleoplasmic localization is supported in HPA's ICC-IF summary, and HPA049480 is ICC Supported (HPA subcellular; HPA antibodies). That observation can guide the separate IF/ICC page; it is not an IHC-P protocol or a substitute for IHC controls. |
| Situation | Likely cause | Next action |
|---|---|---|
| Positive tissue is blank. | A failed stain or an unsuitable run is possible; a High HPA observation does not guarantee positivity in every specimen (HPA tissue IHC: High groups; Approved, medium consistency). | Repeat with an HPA-reported positive cell population and appropriate run controls. Check the catalog antibody's validated IHC-P instructions and detection steps; no target-specific retrieval setting is supplied here. |
| Color is mainly outside nuclei. | The compartment conflicts with the nuclear reference pattern (UniProt Q8IU81: nucleus; HPA tissue IHC: general nuclear expression). Background or cross-reactive binding may be contributing. | Confirm the cell boundaries and nuclear counterstain, then compare reagent controls. Score only interpretable nuclear signal and document persistent discordant staining. |
| Staining is widespread and hazy. | Diffuse background can obscure a nuclear pattern (UniProt Q8IU81: nucleus). In general IHC practice, detection background or overdevelopment may produce broad color. | Inspect control sections and the detection run. Adjust routine blocking, washes, or chromogen development as indicated by those controls, then reassess whether nuclei are individually resolved. |
| Unexpected cells stain strongly. | HPA reports general nuclear expression but identifies particular High cell populations; an unexpected result may reflect true variation, cross-reactivity, or endogenous detection activity (HPA tissue IHC). | Verify the cell type on morphology and compare expected positive cells in the same run. Use suitable detection controls before assigning the unexpected signal to IRF2BP1. |
| Heart cardiomyocytes show convincing nuclear color. | HPA reports IRF2BP1 as Not detected in cardiomyocytes (HPA tissue IHC: heart muscle). This discrepancy merits review, although the HPA profile awaits external verification. | Confirm that the colored nuclei belong to cardiomyocytes, examine background and detection controls, and compare an HPA-reported positive tissue before treating the finding as biological expression. |
| Tonsil germinal-center cells appear faint. | Low staining is reported for that cell population (HPA tissue IHC: tonsil germinal-center cells), so a weak result there alone is a poor basis for declaring the run unsuccessful. | Judge run performance using controls and an HPA-reported High population. Record faint nuclear staining separately from diffuse background rather than forcing a positive-or-negative call. |
Experiment evidence
IHC & IF images showing IRF2BP1 staining
A11406-2 · IHCIHC analysis of IRF2BP1 using anti-IRF2BP1 antibody (A11406-2). IRF2BP1 was detected in a paraffin-embedded section of human acinic cell car
A11406-2 · IHCIHC analysis of IRF2BP1 using anti-IRF2BP1 antibody (A11406-2). IRF2BP1 was detected in a paraffin-embedded section of human acinic cell car
A11406-2 · IFIF analysis of IRF2BP1 using anti-IRF2BP1 antibody (A11406-2). IRF2BP1 was detected in an immunocytochemical section of TPC1 cells. Enzyme a
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000170604-IRF2BP1/tissue/cerebral+cortex#imid_11493115
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000170604-IRF2BP1/tissue/colon#imid_11493049
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000170604-IRF2BP1/tissue/liver#imid_11493042
HPA · PancreasHuman Protein Atlas IHC, Pancreas. Source: https://www.proteinatlas.org/ENSG00000170604-IRF2BP1/tissue/pancreas#imid_11493009
HPA · ICC-IF A-549Human Protein Atlas immunofluorescence, A-549, antibody HPA049480; green = IRF2BP1, blue = nucleus, red = microtubules.
HPA · ICC-IF A-549Human Protein Atlas immunofluorescence, A-549, antibody HPA049480; green = IRF2BP1, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for IRF2BP1 IHC & IF
🧪Run colon first: its glandular cells must show nuclear staining to match the HPA High result (HPA: High in colon glandular cells; UniProt: nucleus). Use heart muscle as the negative tissue (HPA: not detected in cardiomyocytes); cells on the colon slide without detectable staining should show no nuclear signal above background, although HPA does not identify a validated internal negative cell type.
Positive control tissue: Adrenal gland (Glandular cells, HPA High)
Negative control tissue: Heart muscle (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show IRF2BP1 in A-549, HEK293, U2OS, NIH 3T3, with annotated localisation: Nucleoplasm (supported) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control; a concentration-matched rabbit IgG control matched to the primary antibody’s clonality if known (selected-SKU caption: rabbit primary antibody); and IRF2BP1-knockout tissue as a biological negative. Quench endogenous peroxidase and check for background DAB signal in the colon section before scoring glandular nuclei (selected-SKU caption: HRP/DAB detection; HPA: High in colon glandular cells).
⚠️Feasibility: The selected-SKU paraffin-section caption does not report a fixative, and no target-specific fixation window or fixation effect is reported (selected-SKU caption: fixative not stated). Heat-mediated EDTA retrieval at pH 8.0 was used in that IHC example, but whether retrieval is required is unreported (selected-SKU caption: EDTA retrieval). Frozen sections are not established as easier; IF/ICC has supported nucleoplasmic localisation, while colon luminal material may complicate interpretation of glandular staining (HPA: nucleoplasm, supported; HPA: High in colon glandular cells).
HPA tissue IHC evidence for IRF2BP1
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Medium consistency between antibody staining and RNA expression data. Pending external verification.). 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 |
|---|---|---|---|---|
| Adrenal gland | Glandular cells | High | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | High | Protein (IHC) | HPA → |
| Cervix | Squamous epithelial cells | High | Protein (IHC) | HPA → |
| Colon | Glandular cells | High | Protein (IHC) | HPA → |
| Duodenum | Glandular cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Heart muscle | Cardiomyocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced IRF2BP1 IHC Tips
Troubleshoot IRF2BP1 staining by checking nuclear localisation, section processing and cell-level controls before comparing signal intensity (UniProt: nucleus; HPA: general nuclear expression).
How should I adjust retrieval when IRF2BP1 nuclear staining is weak?
Start with heat-mediated retrieval in EDTA at pH 8.0 for the catalog antibody (datasheet A11406-2). The demonstrated paraffin-section workflow used this retrieval before overnight primary incubation at 4°C (caption A11406-2). If nuclear signal remains weak, compare retrieval heating and cooling conditions on adjacent sections while holding antibody concentration and detection constant (standard IHC practice). Assess preserved tissue morphology and nuclear contrast alongside signal, because excess heating can damage sections and make apparent gains difficult to interpret (standard IHC practice). Treat any alternative buffer or pH as a fallback requiring its own controlled comparison (standard IHC practice).
Can fixation explain weak or uneven IRF2BP1 staining in paraffin sections?
Target-specific fixation sensitivity is unknown: the selected paraffin-section caption does not state a fixative (caption A11406-2). Record each specimen’s fixative and fixation interval, then compare similarly processed sections before attributing intensity differences to IRF2BP1 abundance (standard IHC practice). Where processing varies, run the same EDTA pH 8.0 retrieval and 2 μg/ml primary concentration across a small comparison set (caption A11406-2; standard IHC practice). Inspect nuclear morphology, section adhesion and edge-to-centre gradients, since these help identify processing variation (standard IHC practice). Neither the reported tissue staining nor the protein’s nuclear localisation establishes a target-specific fixation effect (HPA: general nuclear expression; UniProt: nucleus).
Should cytoplasmic staining count as an IRF2BP1-positive IHC result?
Prioritise nuclear staining: IRF2BP1 is annotated in the nucleus, and tissue IHC shows a general nuclear pattern (UniProt: nucleus; HPA: general nuclear expression). Supported nucleoplasmic localisation from ICC/IF offers a more specific compartment expectation, although section morphology limits that distinction in chromogenic IHC (HPA: nucleoplasm supported; standard IHC practice). Score nuclear and cytoplasmic signal separately rather than combining both into a single positive call (standard IHC practice). Check that DAB lies within intact, counterstained nuclei and repeats across comparable cells away from damaged edges (standard IHC practice). Predominantly cytoplasmic staining warrants review of background, retrieval and antibody controls before biological interpretation (standard IHC practice).
What can explain discordant staining when the IRF2BP1 epitope is unspecified?
The supplied record lists 0 isoforms and gives no antibody epitope, so an isoform-specific staining claim is unsupported (UniProt: isoform record; caption A11406-2). IRF2BP1 has no annotated transmembrane segment and is assigned to the nucleus; interpret compartment discordance against those observations first (UniProt: topology and subcellular location). The record includes phosphorylated residues and an arginine methylation site, but it does not establish whether either alters this antibody’s binding (UniProt: modified residues; caption A11406-2). Compare adjacent sections with the same retrieval and detection settings, then seek independent specificity evidence if patterns disagree (standard IHC practice). Do not assign an affected residue without an epitope map (standard IHC practice).
How can IF help resolve ambiguous IRF2BP1 staining in tissue?
Use IF as a separate localisation check, pairing IRF2BP1 with a marker identifying the cell population being assessed and a nuclear counterstain (standard IF practice). Expect nuclear, particularly nucleoplasmic, signal rather than membrane-restricted staining (UniProt: nucleus and no transmembrane segment; HPA: nucleoplasm supported). Choose fluorophores after checking tissue autofluorescence, and favour channels with clear separation from that background (standard IF practice). Because the expected epitope is intracellular and nuclear, permeabilise sufficiently for nuclear access while preserving morphology (UniProt: nucleus; standard IF practice). Validate the IF antibody and its fixation conditions independently; the catalog caption documents paraffin-section chromogenic IHC, not IF performance (caption A11406-2; standard IF practice).
How do I distinguish nonspecific DAB deposition from IRF2BP1 staining?
Compare intact nuclear DAB with diffuse cytoplasmic, stromal or edge-associated colour before calling a section positive (UniProt: nucleus; standard IHC practice). The selected example used 10% goat-serum blocking, 2 μg/ml rabbit primary overnight at 4°C, and peroxidase-linked detection with DAB (caption A11406-2). Include a no-primary control and a peroxidase block to assess secondary-reagent binding and endogenous enzyme signal (standard IHC practice). If background persists, titrate primary concentration and detection exposure while keeping retrieval and section processing matched (standard IHC practice). Check whether nuclear signal remains interpretable after background falls, rather than judging specificity from total brown intensity (standard IHC practice).
What scoring method makes IRF2BP1 IHC comparable across sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and score nuclear signal only, consistent with the reported tissue pattern and nuclear annotation (HPA: general nuclear expression; UniProt: nucleus). Record the percentage of positive cells and nuclear intensity categories, then calculate an H-score if graded intensity is reproducible (standard IHC practice). For spatial questions, report positive nuclei per mm² of viable tissue alongside the denominator and sampling area (standard IHC practice). Normalise comparisons to the same cell population, viable area, staining batch and exposure or scan settings (standard IHC practice). Keep cytoplasmic background and excluded necrotic regions documented separately so they cannot inflate the nuclear score (standard IHC practice).
What evidence supports a true IRF2BP1-positive IHC result?
A credible result places DAB in intact nuclei across a coherent cell population, matching the reported general nuclear staining and nuclear annotation (HPA: general nuclear expression; UniProt: nucleus). HPA reports high staining in colon glandular cells and no detection in heart-muscle cardiomyocytes; these are reference patterns, not universal specimen controls (HPA: high in colon glandular cells; HPA: not detected in cardiomyocytes). Exclude isolated tissue-edge signal, necrotic areas and diffuse deposits, and review the no-primary and peroxidase-block controls (standard IHC practice). Interpret discordant cell or compartment staining cautiously, especially because the tissue-IHC profile has medium agreement with RNA data and awaits external verification (HPA: Approved reliability description).
Boster reagents
Best IRF2BP1 / Interferon regulatory factor 2-binding protein 1 IHC Antibodies
A11406-2 has IHC images from human paraffin sections (catalog IHC captions) and an IF image from TPC1 cells (catalog IF caption); listed reactivity covers human, mouse and rat (catalog reactivity).
Real IHC data
Anti-IRF2BP1 Antibody ®
Cat # A11406-2
A11406-2 is listed for IHC and IF/ICC, with human, mouse and rat reactivity (catalog applications and reactivity). Its images show human parotid acinic cell carcinoma and cervical cancer paraffin sections by IHC (catalog IHC captions) and TPC1 cells by IF (catalog IF caption).
Which to pick: For tissue IHC, choose A11406-2: its parotid section image documents EDTA retrieval at pH 8.0, 2 μg/ml primary antibody and HRP/DAB detection; the fixative is unreported (A11406-2 IHC caption). For IF/ICC, the same SKU has a TPC1-cell IF image using 5 μg/ml primary antibody (A11406-2 IF caption). For mouse or rat samples, A11406-2 has listed reactivity, while the supplied IHC and IF images use human samples (catalog reactivity; A11406-2 IHC/IF captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q8IU81 (I2BP1_HUMAN, Interferon regulatory factor 2-binding protein 1).
- Human Protein Atlas. IRF2BP1 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. IRF2BP1 subcellular location (ICC-IF): Localized to the nucleoplasm..
- Human Protein Atlas. IRF2BP1 antibody validation summary (2 antibodies).
- Quantitative proteome profiling of lymph node-positive vs. -negative colorectal carcinomas pinpoints MX1 as a marker for lymph node metastasis. International journal of cancer 2014 — PMC4192020.
- PubMed PMID:12799427 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
- PubMed PMID:17974005 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for IRF2BP1 (Interferon regulatory factor 2-binding protein 1). Expected localisation General nuclear staining observed (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A11406-2). Validated IHC antibody: A11406-2 at 2-5 μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Glandular cells); negative Heart muscle. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human acinic cell carcinoma of parotid cancer tissue; fixative not specified…; 2. Fixation: Image fixative and duration unreported (datasheet A11406-2); 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 retrieval: EDTA pH 8.0 (datasheet A11406-2); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A11406-2); 8. Primary antibody: Rabbit anti-IRF2BP1, 2-5 μg/ml (datasheet A11406-2); 9. Primary incubation: Overnight at 4 °C (datasheet A11406-2). Watch for: Fixation: Keep fixation consistent across sections. (standard IHC practice; not target-specific); Caveat: Staining has medium consistency with RNA expression (HPA tissue IHC); Isoform / epitope: One full-length chain; no isoforms annotated (UniProt); Regulation: No expression regulator annotated (UniProt). Evidence: UniProt Q8IU81, HPA tissue IHC · Approved, HPA ICC-IF · nucleoplasm., Datasheet A11406-2, PMC4192020. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/IRF2BP1/immunohistochemistry-ihc-guide.