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EHD3 / EH domain-containing protein 3 · IHC design guide
Design Immunohistochemistry for EHD3
Plan EHD3 chromogenic IHC on paraffin sections using the catalog antibody's IHC-P conditions (datasheet A04576-1). Assess the cytoplasmic tissue pattern (HPA tissue IHC) while accounting for the warning that the antibody may detect proteins from more than one gene (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
EHD3 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 selected tissue cells (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic in endothelial, trophoblastic and some germinal center cells (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A04576-1) | |
| Positive control | Kidney+4 more · see all | |
| Negative control | Adrenal gland+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A04576-1) | |
| Caveat | Antibody may detect proteins from more than one gene (HPA tissue IHC) | |
| Regulation | Staining regulation unreported (UniProt) | |
| Isoform / epitope | 2 isoforms; no transmembrane segment; epitope coverage unknown (UniProt) |
Section 1
Recommended EHD3 IHC & IF Protocols
The catalog antibody protocol is accompanied by two published chromogenic IHC protocols for EHD3 in rat liver and pituitary adenoma sections (PMC10418749; PMC12715928).
| Sample | Paraffin-embedded mouse brain tissue; fixative not specified (datasheet A04576-1) |
| Fixation | Image fixative and duration unreported (datasheet A04576-1); 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 A04576-1); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A04576-1) |
| Primary antibody | Rabbit anti-EHD3, 2μg/ml (datasheet A04576-1) |
| Primary incubation | Overnight at 4 °C (datasheet A04576-1) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet A04576-1) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | EHD3-positive staining in cells in glomeruli of kidney (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in endothelial, trophoblastic and a subset of germinal center cells. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA at pH 8.0 for the catalog antibody (datasheet: A04576-1); a published protocol used heated citrate for 15 min (PMC12715928).
Section 2
What Is the Expected EHD3 Staining Pattern?
EHD3 associates with recycling endosome, cell and cilium membranes despite having no transmembrane segment (UniProt Q9NZN3). In paraffin IHC, expect cytoplasmic staining in endothelial and trophoblastic cells and a subset of germinal center cells (HPA tissue IHC). Kidney glomerular cells show high staining (HPA tissue IHC). Treat this as a guide to interpretation: HPA rates the tissue pattern Approved, with medium RNA concordance and a warning that the antibodies target proteins from more than one gene (HPA tissue IHC).
| Cytoplasmic stain in endothelial or trophoblastic cells, or a subset of germinal center cells (HPA tissue IHC). | This fits the reported tissue pattern (HPA tissue IHC). Compare the stained cells with the section's anatomy; the pattern supports interpretation but does not prove that every signal is EHD3-specific, because HPA warns of antibodies targeting more than one gene (HPA tissue IHC). |
| Prominent nuclear stain or uniform staining across compartments, with little of the reported cytoplasmic pattern (HPA tissue IHC). | A dominant nuclear pattern lacks support from the reported membrane and recycling endosome locations (UniProt Q9NZN3; HPA subcellular ICC-IF). Check morphology, counterstain and controls before calling it positive; compartment mismatch alone does not identify the source of the signal (general IHC practice). |
| Strong chromogen in cell types listed as not detected, such as adrenal glandular or bronchial respiratory epithelial cells (HPA tissue IHC). | This conflicts with those HPA observations, although a listed negative is not an absolute absence claim (HPA tissue IHC). Consider antibody cross-reactivity, endogenous detection activity or misplaced cell identification; controls and tissue context distinguish these possibilities (general IHC practice). |
| Haze, edge staining or widespread deposit that obscures cell boundaries (general IHC practice). | Score this as background until a cell-associated pattern can be resolved (general IHC practice). Examine a no-primary control and the distribution of deposit; diffuse colour cannot establish the cytoplasmic, cell-selective HPA pattern (HPA tissue IHC). |
| No visible stain in kidney glomerular cells, even though they are an HPA high-staining reference (HPA tissue IHC). | A missing reference signal makes a negative result elsewhere hard to interpret (general IHC practice). First check that glomeruli are present and that detection controls worked; then review the IHC assay settings. HPA's high label describes its observation, not a guarantee for every antibody or section (HPA tissue IHC). |
💡Expected EHD3 appearanceCall the section positive when distinct cytoplasmic chromogen appears in the reported endothelial, trophoblastic or subset of germinal center cells, with kidney glomerular cells a high-staining reference; dominant nuclear colour or uniform staining of reported negative cell types is suspect (HPA tissue IHC; UniProt Q9NZN3).
| Tissue and cell context (HPA tissue IHC). | Kidney glomerular cells are high; placental trophoblastic and lymph node germinal center cells are medium, while heart cardiomyocytes are low in HPA IHC (HPA tissue IHC). UniProt reports high heart and brain tissue expression (UniProt Q9NZN3); that tissue-level statement does not assign a high IHC score to cardiomyocytes or every brain cell. |
| Compartment and resolution (UniProt Q9NZN3; HPA subcellular ICC-IF). | Recycling endosome, cell and cilium membranes are reported for EHD3 (UniProt Q9NZN3). HPA ICC-IF chiefly resolves plasma membrane, with primary cilium and transition-zone signals also supported (HPA subcellular ICC-IF). These fine structures may not be separately resolved in chromogenic tissue IHC (general IHC practice). |
| Antibody evidence and specificity limit (HPA tissue IHC; HPA antibodies). | Two listed antibodies have Approved IHC status, while the tissue summary reports medium consistency with RNA and cautions that antibodies target proteins from more than one gene (HPA antibodies; HPA tissue IHC). An anatomically plausible signal therefore remains an observation requiring appropriate controls, not a unique molecular identification. |
| Isoforms and epitope (UniProt Q9NZN3). | UniProt lists two EHD3 isoforms and an EH domain at residues 444–532 (UniProt Q9NZN3). The supplied antibody record gives no epitope positions or isoform coverage (HPA antibodies). Do not infer which isoform produced a stain, or claim that a particular retrieval condition exposes its epitope. |
| Processing and topology (UniProt Q9NZN3). | The record lists a 1–535 chain, no signal peptide or propeptide, and no transmembrane segment (UniProt Q9NZN3). Interpret membrane-associated staining alongside its reported membrane binding and trafficking role; these annotations provide no basis to expect a shed extracellular staining pattern (UniProt Q9NZN3). |
| Situation | Likely cause | Next action |
|---|---|---|
| Kidney glomeruli appear unstained despite their high HPA IHC level (HPA tissue IHC). | The section may lack evaluable glomeruli, or the assay may have failed; HPA's level is an observed reference rather than a guaranteed result (HPA tissue IHC; general IHC practice). | Confirm the cell population on the counterstained section and verify detection controls. Review the antibody's IHC instructions and the run's retrieval and detection records without assuming EHD3-specific fixation sensitivity (general IHC practice). |
| Colour is diffuse or also appears in a no-primary control (general IHC practice). | Background from the detection workflow can obscure cell-associated stain (general IHC practice). | Compare test and no-primary sections, inspect wash and blocking steps, and adjust the detection workflow according to its controls. Score EHD3 only where the reported cell-selective cytoplasmic pattern remains visible (general IHC practice; HPA tissue IHC). |
| Nuclear stain dominates while the reported cytoplasmic cell pattern is absent (HPA tissue IHC). | The dominant compartment does not match the reported EHD3 locations (UniProt Q9NZN3; HPA subcellular ICC-IF); its cause is unresolved. | Check the counterstain and negative control, then compare staining across the reported positive cell populations. Avoid assigning a nuclear EHD3 location from this slide alone (general IHC practice; HPA tissue IHC). |
| Strong stain appears in HPA not-detected adrenal glandular or bronchial epithelial cells (HPA tissue IHC). | Possible explanations include cross-reactivity, endogenous detection activity or cell identification error; the HPA antibody warning makes specificity a concern (HPA tissue IHC; general IHC practice). | Recheck anatomy and run an appropriate no-primary and detection control. Report the discrepancy rather than treating the unexpected cell population as confirmed EHD3 expression (general IHC practice; HPA tissue IHC). |
| Heart cardiomyocytes stain weakly although UniProt reports high expression in heart (UniProt Q9NZN3). | HPA specifically lists cardiomyocytes as low by tissue IHC; whole-tissue expression and cell-level stain intensity are different observations (HPA tissue IHC; UniProt Q9NZN3). | Use the HPA cardiomyocyte observation when judging this IHC pattern. Confirm assay performance with a reported high-staining population before interpreting absent cardiac signal (HPA tissue IHC; general IHC practice). |
| IF/ICC Q: Should a plasma-membrane or cilium signal be judged by the paraffin IHC pattern? | HPA's membrane, cilium and transition-zone localisation comes from ICC-IF; its tissue IHC summary describes cytoplasmic cell patterns (HPA subcellular ICC-IF; HPA tissue IHC). | A: Interpret IF/ICC against its own localisation evidence and controls; do not require the same visual resolution in chromogenic paraffin IHC. HPA flags multi-gene targeting for the subcellular result, so localisation alone does not settle specificity (HPA subcellular ICC-IF; general IHC practice). |
Experiment evidence
IHC & IF images showing EHD3 staining
A04576-1 · IHCIHC analysis of EHD3 using anti-EHD3 antibody (A04576-1). EHD3 was detected in paraffin-embedded section of mouse brain tissue. Heat mediate
A04576-1 · IHCIHC analysis of EHD3 using anti-EHD3 antibody (A04576-1). EHD3 was detected in paraffin-embedded section of human ovarian serous adenocarcin
A04576-1 · IFIF analysis of EHD3 using anti-EHD3 antibody (A04576-1). EHD3 was detected in immunocytochemical section of MCF-7 cells. Enzyme antigen retr
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000013016-EHD3/tissue/cerebral+cortex#imid_15783183
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000013016-EHD3/tissue/colon#imid_15772749
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000013016-EHD3/tissue/liver#imid_15772750
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000013016-EHD3/tissue/kidney#imid_15772743
![HPA ICC-IF BJ [Human fibroblast] — EHD3 (HPA066751)](https://images.proteinatlas.org/66751/1667_E12_5_blue_red_green.jpg)
HPA · ICC-IF BJ [Human fibroblast]Human Protein Atlas immunofluorescence, BJ [Human fibroblast], antibody HPA066751; green = EHD3, blue = nucleus, red = microtubules.
![HPA ICC-IF BJ [Human fibroblast] — EHD3 (HPA066751)](https://images.proteinatlas.org/66751/1667_E12_6_blue_red_green.jpg)
HPA · ICC-IF BJ [Human fibroblast]Human Protein Atlas immunofluorescence, BJ [Human fibroblast], antibody HPA066751; green = EHD3, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for EHD3 IHC & IF
🧪Run kidney first: cells in glomeruli should stain (HPA: High in cells in glomeruli). Use adrenal gland as the negative tissue: glandular cells are reported as not detected (HPA: Not detected in adrenal gland glandular cells). On the kidney slide, internal negative cells should remain at background, but identify them by independent validation because the HPA kidney row does not establish which nonglomerular cells lack EHD3 (HPA: High in cells in glomeruli).
Positive control tissue: Kidney (Cells in glomeruli, HPA High)
Negative control tissue: Adrenal gland (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show EHD3 in BJ [Human fibroblast], SiHa, U2OS, ASC52telo, RPTEC/TERT1, hTERT-RPE1 (serum starved), KOLF2.1J, with annotated localisation: Plasma membrane (supported) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control; a concentration-matched rabbit IgG control matched to the primary’s clonality; and EHD3 knockout tissue as a biological negative (caption: rabbit anti-EHD3 primary; standard IHC practice). For kidney, quench endogenous peroxidase and check for endogenous biotin background with the caption’s biotinylated secondary and SABC–DAB detection (caption: biotinylated secondary/SABC/DAB; standard IHC practice).
⚠️Feasibility: No EHD3-specific fixation window or fixation effect is reported, and the A04576-1 paraffin-section caption does not state a fixative (caption: fixative unreported). The caption uses heat retrieval in EDTA at pH 8.0, but does not establish that retrieval is required; paraffin IHC is directly documented, while the supplied IF evidence consists of cell-line ICC-IF images and does not establish that frozen sections or IF are easier (caption: EDTA retrieval; HPA: ICC-IF cell-line images). Kidney staining with the caption’s biotin-based DAB method needs a background check for endogenous biotin (caption: SABC/DAB; standard IHC practice).
HPA tissue IHC evidence for EHD3
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Medium consistency between antibody staining and RNA expression data. Caution, targets protein from more than one gene. 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 |
|---|---|---|---|---|
| Kidney | Cells in glomeruli | High | Protein (IHC) | HPA → |
| Adipose tissue | Adipocytes | Medium | Protein (IHC) | HPA → |
| Appendix | Lymphoid tissue | Medium | Protein (IHC) | HPA → |
| Lung | Alveolar cells | Medium | Protein (IHC) | HPA → |
| Lymph node | Germinal center cells | Medium | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adrenal gland | 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 → |
| Cerebellum | Cells in granular layer | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced EHD3 IHC Tips
Troubleshoot EHD3 staining in paraffin sections by checking retrieval, cellular context and controls before interpreting chromogenic signal.
Which retrieval conditions should I try first for weak EHD3 staining?
Start with heat mediated antigen retrieval in EDTA at pH 8.0 for paraffin sections (datasheet A04576-1). The selected mouse brain IHC example then used 2 μg/mL catalog antibody overnight at 4°C, providing a matched starting condition for troubleshooting (datasheet A04576-1). If staining remains weak, vary heating time in small increments while keeping section thickness, antibody concentration and detection constant (standard IHC practice). Compare each change with a matched positive section and a no primary control, because stronger DAB alone cannot establish EHD3 specificity (standard IHC practice). Record tissue integrity alongside signal, since excessive heating can damage morphology (standard IHC practice).
Could fixation explain weak or uneven EHD3 staining?
Target specific fixation sensitivity is unknown: the selected paraffin section caption does not state a fixative (datasheet A04576-1). Document the fixative, fixation duration and tissue processing for each specimen before comparing DAB intensity across cases (standard IHC practice). If staining varies between batches, process replicate sections with the same EDTA pH 8.0 retrieval and detection conditions to isolate processing differences (datasheet A04576-1; standard IHC practice). Avoid calling a poorly stained section EHD3 negative until a positive control processed in parallel stains acceptably (standard IHC practice). Do not infer a fixation effect from tissue expression patterns or EHD3 sequence features (HPA tissue IHC; UniProt Q9NZN3).
Where should convincing EHD3 signal appear in paraffin sections?
Assess cellular and membrane associated cytoplasmic staining in the relevant cells, consistent with EHD3 at recycling endosome and cell membranes (UniProt Q9NZN3 subcellular location). A ciliary pocket signal may be plausible where that structure can be resolved, but routine DAB sections rarely establish its identity without an independent marker (UniProt Q9NZN3 subcellular location; standard IHC practice). HPA describes cytoplasmic staining in endothelial, trophoblastic and some germinal center cells, while reporting high signal in cells of kidney glomeruli (HPA tissue IHC). Compare candidate staining with tissue anatomy and a matched negative control before assigning it to those cells (standard IHC practice). Diffuse nuclear staining alone conflicts with the supplied localisation evidence (UniProt Q9NZN3 subcellular location).
Can this antibody distinguish EHD3 isoforms or modified epitopes?
The record lists 2 EHD3 isoforms, but the supplied caption does not identify the antibody epitope or establish isoform discrimination (UniProt Q9NZN3 isoforms; datasheet A04576-1). EHD3 contains a dynamin type G domain at residues 55–286 and an EH domain at 444–532; their presence does not locate this antibody’s binding site (UniProt Q9NZN3 domains). Phosphoserines at residues 349 and 456 are recorded, without evidence that they alter this IHC stain (UniProt Q9NZN3 modified residues). For an isoform or modification claim, obtain the catalog antibody’s immunogen information and validate against an independent, appropriately controlled assay (standard IHC practice). Score the present DAB signal as antibody reactivity, not a specific EHD3 isoform (standard IHC practice).
How can IF help resolve ambiguous EHD3 positive cells?
Use IF on a separately validated specimen to pair EHD3 with a marker of the proposed cell type, such as CD31 when evaluating endothelial cells (HPA tissue IHC; standard IF practice). Choose a far red fluorophore when tissue autofluorescence obscures shorter wavelength channels, and inspect single stain controls before interpreting overlap (standard IF practice). Because EHD3 has no transmembrane segment and is associated with recycling endosome and cell membranes, optimise permeabilisation for access to the antibody’s epitope without assuming which side it faces (UniProt Q9NZN3 topology and subcellular location; standard IF practice). Confirm the catalog antibody’s suitability for IF independently; its selected image documents paraffin section IHC with DAB (datasheet A04576-1). Keep IF colocalisation conclusions separate from chromogenic IHC scores (standard IF practice).
How do I distinguish EHD3 signal from chromogenic background?
Run a no primary control and examine whether brown deposit follows cells or instead accumulates at folds, cut edges and damaged areas (standard IHC practice). In the selected mouse brain example, sections were blocked with 10% goat serum and developed through a biotin based system with DAB (datasheet A04576-1). Check endogenous peroxidase blocking and, for biotin based detection, endogenous biotin controls before attributing widespread brown signal to EHD3 (standard IHC practice). If background persists, titrate the primary antibody around the documented 2 μg/mL condition while holding retrieval and exposure constant (datasheet A04576-1; standard IHC practice). Judge improvement by cellular contrast and control staining, not total darkness (standard IHC practice).
What should an EHD3 IHC score measure and use as its denominator? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, then use an H-score for intensity weighted positive cells or report the percentage of positive cells (standard IHC practice). For spatial questions, report EHD3 positive cell density per mm² of viable tissue and state the analysed area (standard IHC practice). Normalise cell based scores to the number of eligible cells in the same annotated compartment, rather than all cells across unlike tissue regions (standard IHC practice). Keep retrieval, DAB development and image thresholds consistent across specimens, and exclude folds, necrosis and edges using preset rules (standard IHC practice). Report the scoring method alongside the cellular pattern, since EHD3 staining varies by cell population (HPA tissue IHC; standard IHC practice).
When is an apparent EHD3 positive result credible?
A credible result combines anatomically plausible cellular staining with a clean no primary control and a working positive control (standard IHC practice). EHD3 is reported at recycling endosome and cell membranes, while HPA describes cytoplasmic staining in endothelial, trophoblastic and some germinal center cells (UniProt Q9NZN3 subcellular location; HPA tissue IHC). Question isolated nuclear signal, staining confined to cut edges or necrosis, and brown precipitate that persists without primary antibody (UniProt Q9NZN3 subcellular location; standard IHC practice). HPA rates its tissue staining as Approved but cautions that the antibody targets proteins from more than one gene and awaits external verification (HPA tissue IHC). Treat a cell identity or EHD3 specific claim as provisional until an independent control supports it (standard IHC practice).
Boster reagents
Best EHD3 / EH domain-containing protein 3 IHC Antibodies
The IHC-validated anti-EHD3 antibody has paraffin-section images from mouse brain and human ovarian and renal tumors, plus an IF/ICC image from MCF-7 cells (catalog image captions).
Real IHC data
Anti-EHD3 Antibody ®
Cat # A04576-1
A04576-1 has IHC images from paraffin-embedded mouse brain, human ovarian serous adenocarcinoma, and human renal clear cell carcinoma (A04576-1 IHC captions). A04576-1 is also listed for IF/ICC and has an IF image from MCF-7 cells; its listed reactivity is human, monkey, mouse, and rat (catalog applications, IF caption, and reactivity).
Which to pick: Choose A04576-1 for tissue IHC when its paraffin-section examples match your workflow: the captions describe EDTA retrieval at pH 8.0 and 2 µg/mL primary antibody, but do not report the fixative (A04576-1 IHC captions). The same SKU is listed for IF/ICC and has an MCF-7 IF image; its listed reactivity spans human, monkey, mouse, and rat, while the supplied IHC images show only human and mouse tissue (catalog applications, reactivity, and image captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q9NZN3 (EHD3_HUMAN, EH domain-containing protein 3).
- Human Protein Atlas. EHD3 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. EHD3 subcellular location (ICC-IF): Mainly localized to the plasma membrane. In addition localized to the primary cilium and primary cilium transition zone. Caution: Based on antibodies targeting proteins from multiple genes..
- Human Protein Atlas. EHD3 antibody validation summary (3 antibodies).
- Proteomic Analysis of Dysfunctional Liver Sinusoidal Endothelial Cells Reveals Substantial Differences in Most Common Experimental Models of Chronic Liver Diseases. International journal of molecular sciences 2023 — PMC10418749.
- Revealing sphingolipid metabolism genes as biomarkers for the diagnosis of invasive pituitary adenomas in silico and in vivo. European journal of medical research 2025 — PMC12715928.
- Engineering scalable vascularized kidney organoids for in vivo glomerular filtration with human endothelial integration. npj biomedical innovations 2026 — PMC12864042.
- PubMed PMID:10673336 — UniProt-cited evidence.
- PubMed PMID:12121420 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for EHD3 (EH domain-containing protein 3). Expected localisation Cytoplasmic staining in selected tissue cells (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A04576-1). Validated IHC antibody: A04576-1 at 2μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Kidney (Cells in glomeruli); negative Adrenal gland. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded mouse brain tissue; fixative not specified (datasheet A04576-1); 2. Fixation: Image fixative and duration unreported (datasheet A04576-1); 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 A04576-1); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A04576-1); 8. Primary antibody: Rabbit anti-EHD3, 2μg/ml (datasheet A04576-1); 9. Primary incubation: Overnight at 4 °C (datasheet A04576-1). Watch for: Fixation: Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A04576-1); Caveat: Antibody may detect proteins from more than one gene (HPA tissue IHC); Isoform / epitope: 2 isoforms; no transmembrane segment; epitope coverage unknown (UniProt); Regulation: Staining regulation unreported (UniProt). Evidence: UniProt Q9NZN3, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the plasma membrane. In addition localized to…, Datasheet A04576-1, PMC10418749, PMC12715928. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/EHD3/immunohistochemistry-ihc-guide.