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INF2 / Inverted formin-2 · IHC design guide
Design Immunohistochemistry for INF2
Plan INF2 IHC in paraffin sections around cytoplasmic staining in most tissues (HPA tissue IHC). Start with the catalog antibody’s 2–5 μg/mL IHC range (datasheet A02710-2) and compare cell-specific staining with controls.
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
INF2 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 | Cytoplasmic staining in most tissues (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic staining across most tissues (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A02710-2) | |
| Positive control | Appendix+4 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 paraffin sections. (standard IHC practice; not target-specific) | |
| Caveat | Staining has medium consistency with RNA expression (HPA tissue IHC) | |
| Regulation | Low tissue specificity (HPA tissue RNA) | |
| Isoform / epitope | 3 isoforms; check whether the epitope is shared (UniProt) |
Section 1
Recommended INF2 IHC & IF Protocols
The catalog antibody uses EDTA pH 8.0 retrieval (datasheet A02710-2). The published IHC protocols below cover glioblastoma, INF2 morphants, and human colorectal tissue (PMC7391617; PMC4457406; PMC12128233).
| Sample | Paraffin-embedded human colon cancer tissue; fixative not specified (datasheet A02710-2) |
| Fixation | Image fixative and duration unreported (datasheet A02710-2); 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 A02710-2); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A02710-2) |
| Primary antibody | Rabbit anti-INF2, 2-5μg/ml (datasheet A02710-2) |
| Primary incubation | Overnight at 4 °C (datasheet A02710-2) |
| Detection | HRP-conjugated secondary, DAB chromogen (datasheet A02710-2) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | INF2-positive staining in glandular cells of appendix (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in most tissues. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA pH 8.0 retrieval for the catalog antibody (datasheet A02710-2); use each paper’s reported retrieval when reproducing its protocol (PMC7391617; PMC4457406; PMC12128233).
Section 2
What Is the Expected INF2 Staining Pattern?
INF2 should appear mainly in the cytoplasm of many tissue cell types, with a perinuclear distribution also consistent with its annotated location (HPA tissue IHC: cytoplasmic expression in most tissues; UniProt Q27J81: cytoplasm, perinuclear region). Glandular cells in appendix, gallbladder, rectum and stomach, and placental trophoblastic cells, show high staining (HPA tissue IHC: High). INF2 has no transmembrane segment (UniProt Q27J81 topology). HPA rates tissue IHC reliability Approved, with medium consistency against RNA expression (HPA tissue IHC).
| Cytoplasmic, possibly perinuclear, staining in appendix glandular cells or placental trophoblastic cells. | This fits INF2 localization and the reported high staining in those cells (UniProt Q27J81: cytoplasm, perinuclear region; HPA tissue IHC: High). Compare signal within the named cell population; staining elsewhere in the section alone is a less direct check of the reported pattern. |
| Predominantly nuclear-only or crisp membrane-only staining in an otherwise positive tissue. | This departs from the reported tissue IHC cytoplasmic pattern and UniProt location (HPA tissue IHC; UniProt Q27J81). Check the control section and detection conditions before assigning the signal to INF2. HPA reports nuclear bodies in ICC-IF, so isolated nuclear puncta need separate interpretation (HPA subcellular ICC-IF). |
| Strong staining of adipocytes, cardiomyocytes or skeletal myocytes, especially without the expected positive cells. | HPA reports these cell types as not detected (HPA tissue IHC). Consider cross-reactivity or endogenous detection activity and compare with an appropriate reagent control (general IHC practice). The HPA observation is a reference pattern, not proof that every positive cell is false. |
| Uniform brown haze across cells and extracellular areas, obscuring cell boundaries. | Diffuse background limits a compartment-level call. Review blocking, washing and detection controls for nonspecific signal or endogenous activity (general chromogenic IHC practice). Do not score the haze as cytoplasmic INF2 solely because HPA describes broad tissue expression (HPA tissue IHC). |
| No staining in appendix glandular cells or placental trophoblastic cells on an otherwise interpretable section. | That conflicts with HPA's high-staining examples (HPA tissue IHC). Check whether the IHC-validated antibody and detection run yield a signal in a positive control, then review retrieval and reagent performance (general IHC practice). Absence in one section does not establish absent INF2 protein. |
💡Expected INF2 appearanceA convincing positive is cell-associated cytoplasmic staining, potentially perinuclear, with high signal in the named glandular or trophoblastic populations (UniProt Q27J81; HPA tissue IHC: High); broad nuclear-only, membrane-only or tissue-wide haze warrants control-based review (general IHC interpretation).
| Tissue and cell selection | Appendix, gallbladder, rectum and stomach glandular cells and placental trophoblastic cells are high-staining examples; adipocytes and cardiomyocytes are reported not detected (HPA tissue IHC). Score the specified cells, since a whole-section average can conceal their pattern. |
| Compartment and topology | UniProt places INF2 in cytoplasm and the perinuclear region and lists no transmembrane segment (UniProt Q27J81). HPA tissue IHC describes cytoplasmic expression in most tissues (HPA tissue IHC). These annotations support a cellular pattern check, not a precise organelle assignment by chromogenic IHC. |
| Antibody evidence | HPA000724 is Approved for IHC, while the tissue profile has medium consistency with RNA data (HPA antibodies; HPA tissue IHC). Interpret an unexpected cell-specific result alongside controls; Approved does not make every staining event target-specific. |
| Isoforms and epitope | UniProt lists three INF2 isoforms and domains at residues 2–330, 554–946 and 974–989 (UniProt Q27J81). No antibody epitope or isoform coverage is supplied, so a difference in staining cannot be assigned to an isoform from this record. |
| IF/ICC Q: What pattern is reported? | A: HPA reports mainly endoplasmic reticulum localization, with additional nuclear bodies, in ICC-IF (HPA subcellular ICC-IF). HPA000724 has Uncertain ICC validation (HPA antibodies). This IF observation can inform interpretation but is not an IHC organelle-level claim. |
| Situation | Likely cause | Next action |
|---|---|---|
| A high-staining reference cell population is blank. | Possible run-level loss of detectable signal; HPA reports high staining in appendix glandular cells and placental trophoblastic cells (HPA tissue IHC). | Inspect the positive control and detection run, then review the antibody's IHC retrieval conditions and reagent performance (general IHC practice). Record the result as unresolved until the control works. |
| Most cells show similar brown haze. | Background may obscure the HPA-reported cytoplasmic pattern (HPA tissue IHC); nonspecific binding or endogenous detection activity are general IHC possibilities. | Compare reagent controls, blocking and wash conditions, and the distribution of signal outside cells (general chromogenic IHC practice). Score defined cells only after background permits that distinction. |
| Signal is sharply membrane-bound. | A membrane-dominant result conflicts with cytoplasmic tissue staining and the absence of a transmembrane segment (HPA tissue IHC; UniProt Q27J81 topology). | Check localization in a HPA high-staining reference population and review detection controls (HPA tissue IHC; general IHC practice). Treat persistent membrane-only staining as unconfirmed. |
| Adipocytes or muscle cells stain more strongly than nearby reference cells. | HPA reports adipocytes, cardiomyocytes and skeletal myocytes as not detected (HPA tissue IHC); cross-reactivity or endogenous activity is possible (general IHC practice). | Compare the named cell types with a positive reference section and reagent controls before calling the unexpected staining INF2 (HPA tissue IHC; general IHC practice). |
| Scoring varies greatly between regions of one section. | INF2 staining is cell-type dependent in the supplied tissue examples: some glandular cells are High, while other listed populations are Low or Not detected (HPA tissue IHC). | Identify and score the relevant cell population in each region rather than averaging all tissue elements; document the observed compartment and intensity (HPA tissue IHC; general IHC practice). |
| ICC-IF shows nuclear puncta, but tissue IHC is mainly cytoplasmic. | HPA lists nuclear bodies as an additional ICC-IF location and cytoplasmic expression as the tissue IHC profile (HPA subcellular ICC-IF; HPA tissue IHC). | Interpret each assay against its own HPA observation. HPA000724 has Uncertain ICC validation, so verify an ICC-IF punctate result independently before relying on it (HPA antibodies). |
Experiment evidence
IHC & IF images showing INF2 staining
A02710-2 · IHCIHC analysis of INF2 using anti-INF2 antibody (A02710-2). INF2 was detected in a paraffin-embedded section of human colon cancer tissue. Hea
A02710-2 · IHCIHC analysis of INF2 using anti-INF2 antibody (A02710-2). INF2 was detected in a paraffin-embedded section of human thyroid cancer tissue. H
A02710-2 · IFIF analysis of INF2 using anti-INF2 antibody (A02710-2). INF2 was detected in an immunocytochemical section of A549 cells. Enzyme antigen re
A02710-1 · IFIF analysis of INF2 using anti-INF2 antibody (A02710-1). INF2 was detected in an immunocytochemical section of A549 cells. Enzyme antigen re
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000203485-INF2/tissue/cerebral+cortex#imid_230770
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000203485-INF2/tissue/colon#imid_230688
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000203485-INF2/tissue/liver#imid_230689
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000203485-INF2/tissue/kidney#imid_230698
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA000724; green = INF2, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA000724; green = INF2, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for INF2 IHC & IF
🧪Run appendix first: glandular cells should stain for INF2 (HPA: High in appendix glandular cells). Run adipose tissue as a negative comparator because adipocytes are listed as not detected (HPA: Not detected in adipocytes); on the appendix slide, assess adjacent nonglandular cells for background staining without assuming they lack INF2.
Positive control tissue: Appendix (Glandular cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show INF2 in A-431, U-251MG, U2OS, with annotated localisation: Endoplasmic reticulum (approved) (HPA subcellular).
Technical controls: Include a no-primary (secondary-only) slide, a rabbit IgG isotype control matched to the catalog antibody’s clonality, and an INF2 knockout sample or validated peptide-block control (caption: rabbit anti-INF2 antibody; standard IHC practice). For chromogenic appendix sections, quench endogenous peroxidase and check for background DAB signal in the controls (caption: HRP/DAB detection; standard IHC practice).
⚠️Feasibility: A target-specific fixation window and fixation effect are unreported; the selected A02710-2 paraffin-section caption does not state a fixative. Its reported IHC conditions use heat retrieval in EDTA at pH 8.0, so test retrieval under those conditions before scoring (caption: EDTA pH 8.0). The supplied evidence does not establish whether frozen sections or IF are easier; in appendix, distinguish glandular cell staining from luminal DAB deposit and peroxidase background using the controls (HPA: High in appendix glandular cells; caption: HRP/DAB detection; standard IHC practice).
HPA tissue IHC evidence for INF2
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Medium consistency between antibody staining and RNA expression data.). 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 |
|---|---|---|---|---|
| Appendix | Glandular cells | High | Protein (IHC) | HPA → |
| Gallbladder | Glandular cells | High | Protein (IHC) | HPA → |
| Placenta | Trophoblastic cells | High | Protein (IHC) | HPA → |
| Rectum | Glandular cells | High | Protein (IHC) | HPA → |
| Stomach | Glandular cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Heart muscle | Cardiomyocytes | Not detected | Protein (IHC) | HPA → |
| Ovary | Ovarian stroma cells | Not detected | Protein (IHC) | HPA → |
| Skeletal muscle | Myocytes | Not detected | Protein (IHC) | HPA → |
| Smooth muscle | Smooth muscle cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced INF2 IHC Tips
Troubleshoot INF2 staining in paraffin sections by checking retrieval, cellular localisation and controls before comparing staining intensity across samples.
Which retrieval condition should I start with for INF2 paraffin IHC?
Start with heat-mediated retrieval in EDTA at pH 8.0 for INF2 paraffin sections (datasheet A02710-2). The selected tissue image used that condition before incubation with 2 μg/mL primary antibody overnight at 4°C, so keep those steps consistent while assessing retrieval (datasheet A02710-2). If staining is weak, compare a modest change in heating duration on adjacent sections while keeping buffer and detection constant, and watch for tissue damage (standard IHC practice). Include a known positive section in each run; strong cytoplasmic staining in glandular cells is a useful reference, although HPA reports only medium consistency between staining and RNA expression (HPA: high in appendix glandular cells; HPA: Approved, medium consistency).
How should I troubleshoot weak INF2 staining when fixation history varies?
The selected paraffin-section caption does not state a fixative, so INF2 sensitivity to fixation cannot be determined from it (datasheet A02710-2). Record the fixative and fixation duration for each specimen, then compare sections processed and stained together before attributing intensity differences to biology (standard IHC practice). Keep EDTA retrieval at pH 8.0 and the 2 μg/mL primary incubation constant during that comparison (datasheet A02710-2). If a poorly staining specimen also has weak staining for an appropriate independent tissue control, investigate processing or antigen preservation; HPA tissue patterns and INF2 topology do not establish a target-specific fixation effect (standard IHC practice; HPA: tissue IHC; UniProt Q27J81: topology).
Where should convincing INF2 staining appear in tissue sections?
Expect predominantly cytoplasmic staining when assessing INF2 by chromogenic IHC; perinuclear enrichment can be consistent with the UniProt annotation (HPA: cytoplasmic expression in most tissues; UniProt Q27J81: cytoplasm, perinuclear region). HPA cell imaging places INF2 mainly at the endoplasmic reticulum and additionally in nuclear bodies, but that higher-resolution observation need not appear as distinct structures in a DAB section (HPA: approved subcellular locations; standard IHC practice). Score staining within identified cells instead of treating an isolated dark nuclear profile as sufficient evidence of INF2 (standard IHC practice). Check whether the distribution is reproducible across intact tissue areas and appropriate positive controls before interpreting unusual compartment staining (standard IHC practice).
Could an INF2 isoform or masked epitope explain discrepant IHC staining?
INF2 has 3 annotated isoforms, and its GBD/FH3, FH2 and WH2 regions occupy different parts of the protein (UniProt Q27J81: isoforms and domains). The supplied caption does not identify the A02710-2 epitope, so staining with this antibody cannot establish which isoform is present (datasheet A02710-2). If specimens disagree, first standardise EDTA retrieval at pH 8.0 and the 2 μg/mL primary incubation used for the selected paraffin section (datasheet A02710-2). For an isoform-specific conclusion, obtain the antibody's epitope mapping and validate it against an independent isoform-resolving assay; localisation or staining intensity alone is insufficient (standard IHC validation practice).
How can IF help resolve ambiguous INF2 staining seen by IHC?
Use IF as a separate validation experiment and multiplex INF2 with a marker identifying the cell population under study; podocytes or tubule cells are relevant kidney populations (UniProt Q27J81: kidney expression; standard IF practice). Choose spectrally separated fluorophores and inspect unstained tissue for autofluorescence before interpreting faint signal (standard IF practice). INF2 has no transmembrane segment and is annotated in cytoplasmic and perinuclear regions, while HPA imaging also supports endoplasmic reticulum localisation (UniProt Q27J81: topology and localisation; HPA: approved endoplasmic reticulum location). Permeabilise sufficiently to expose intracellular antigen, then optimise the IF conditions independently because the supplied 2 μg/mL incubation documents paraffin IHC rather than IF (standard IF practice; datasheet A02710-2).
What should I change when INF2 DAB staining is diffuse or patchy?
First inspect a no-primary section to locate signal arising from the detection system, then review the peroxidase block and DAB development as general chromogenic IHC steps (standard IHC practice). The selected caption reports 10% goat-serum blocking, 2 μg/mL primary overnight at 4°C, and a peroxidase-conjugated secondary for 30 minutes at 37°C (datasheet A02710-2). If diffuse colour persists, compare primary concentration or incubation duration on adjacent sections while holding retrieval and development constant (standard IHC practice). Evaluate background against cell morphology and the expected predominantly cytoplasmic pattern, since widespread colour outside intact cells does not establish INF2 expression (HPA: cytoplasmic expression in most tissues; standard IHC practice).
How should I quantify INF2 staining across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and tissue area before scoring, then record the percentage of positive cells and staining intensity or calculate an H-score using one prespecified scheme (standard IHC quantification practice). For cellular comparisons, normalise positive counts to the number of evaluable cells of that type; for spatial comparisons, report positive-cell density per mm² of viable, analysable tissue (standard IHC quantification practice). Use identical retrieval, antibody incubation, DAB development and image settings across sections to limit technical differences (standard IHC practice). Report compartment and cell type alongside each score because HPA describes cytoplasmic expression across most tissues but also reports varying cell-specific levels (HPA: tissue IHC profile and listed cell levels).
How do I distinguish true INF2 positivity from staining artefact?
Prioritise staining in intact cells with a predominantly cytoplasmic pattern, allowing perinuclear enrichment, and compare it with the negative control and adjacent tissue architecture (HPA: cytoplasmic expression in most tissues; UniProt Q27J81: perinuclear region; standard IHC practice). In kidney, podocytes and some tubule cells are plausible positive populations, but morphology alone cannot confirm antibody specificity (UniProt Q27J81: kidney expression; standard IHC validation practice). Treat staining confined to section edges, necrotic regions or a no-primary control as suspect, and check endogenous peroxidase activity before assigning DAB colour to INF2 (standard IHC practice). HPA's Approved tissue profile has medium consistency with RNA expression, so require reproducible localisation and suitable controls before making a strong biological claim (HPA: Approved, medium consistency).
Boster reagents
Best INF2 / Inverted formin-2 IHC Antibodies
Anti-INF2 antibodies have IHC images from human colon and thyroid cancer and mouse and rat brain paraffin sections (A02710-2 captions), plus A549 cell IF images (A02710-1 and A02710-2 captions).
Real IHC data
Anti-INF2 Antibody ®
Cat # A02710-2
Real IF data
Anti-INF2 Antibody ®
Cat # A02710-1
A02710-2 has paraffin-section IHC images from human colon and thyroid cancer and mouse and rat brain, plus an A549 cell IF image (A02710-2 captions). A02710-1 lists IHC and IF/ICC applications and has an A549 cell IF image; its IHC image is unreported (A02710-1 catalog and caption).
Which to pick: Choose A02710-2 for tissue IHC because its own captions document paraffin sections; the fixative is unreported (A02710-2 IHC captions). For IF/ICC, either SKU has an A549 cell image, and both list IF/ICC; A02710-2 is listed as polyclonal, while A02710-1 clonality is unreported (catalog and IF captions). For work across species, both list human, mouse and rat reactivity, while A02710-2 additionally has IHC images in all three species (catalog and A02710-2 IHC captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q27J81 (INF2_HUMAN, Inverted formin-2).
- Human Protein Atlas. INF2 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. INF2 subcellular location (ICC-IF): Mainly localized to the endoplasmic reticulum. In addition localized to the nuclear bodies..
- Human Protein Atlas. INF2 antibody validation summary (1 antibodies).
- Formin Proteins FHOD1 and INF2 in Triple-Negative Breast Cancer: Association With Basal Markers and Functional Activities. Breast cancer : basic and clinical research 2018 — PMC6109849.
- Multiple formin proteins participate in glioblastoma migration. BMC cancer 2020 — PMC7391617.
- Human Kidney Disease-causing INF2 Mutations Perturb Rho/Dia Signaling in the Glomerulus. EBioMedicine 2014 — PMC4457406.
- Targeting INF2 with DiosMetin 7-O-β-D-Glucuronide: a new stratagem for colorectal cancer therapy. BMC cancer 2025 — PMC12128233.
- PubMed PMID:14702039 — UniProt-cited evidence.
- PubMed PMID:12508121 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for INF2 (Inverted formin-2). Expected localisation Cytoplasmic staining in most tissues (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A02710-2). Validated IHC antibody: A02710-2 at 2-5μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Appendix (Glandular cells); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human colon cancer tissue; fixative not specified (datasheet A02710-2); 2. Fixation: Image fixative and duration unreported (datasheet A02710-2); 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 A02710-2); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A02710-2); 8. Primary antibody: Rabbit anti-INF2, 2-5μg/ml (datasheet A02710-2); 9. Primary incubation: Overnight at 4 °C (datasheet A02710-2). Watch for: Fixation: Keep fixation consistent across paraffin sections. (standard IHC practice; not target-specific); Caveat: Staining has medium consistency with RNA expression (HPA tissue IHC); Isoform / epitope: 3 isoforms; check whether the epitope is shared (UniProt); Regulation: Low tissue specificity (HPA tissue RNA). Evidence: UniProt Q27J81, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the endoplasmic reticulum. In addition localized…, Datasheet A02710-2, PMC7391617, PMC4457406. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/INF2/immunohistochemistry-ihc-guide.