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EIF4A2 / Eukaryotic initiation factor 4A-II · IHC design guide
Design Immunohistochemistry for EIF4A2
Plan EIF4A2 chromogenic IHC on paraffin sections using the reported broad cytoplasmic pattern (HPA tissue IHC). Compare staining across consistently fixed sections and interpret intensity cautiously because the tissue IHC evidence has uncertain reliability (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
EIF4A2 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 | Ubiquitous cytoplasmic staining (HPA tissue IHC) | |
| Staining pattern | Broad cytoplasmic staining, including glandular and glial cells (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet PB9758) | |
| Positive control | Adipose tissue+4 more · see all | |
| Negative control | None in HPA (detected in all 45 tissues); use no-primary + isotype controls |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent across sections. (standard IHC practice; not target-specific) | |
| Caveat | Tissue staining has uncertain reliability; verify specificity (HPA tissue IHC) | |
| Regulation | No specific expression regulator annotated (UniProt) | |
| Isoform / epitope | Two isoforms; epitope coverage is unreported (UniProt; datasheet PB9758) |
Section 1
Recommended EIF4A2 IHC & IF Protocols
The catalog antibody uses EDTA pH 8.0 retrieval (datasheet: PB9758). The published IHC protocols below cover esophageal, colorectal, and FFPE sections (PMC7471732; PMC6518650; PMC13366719).
| Sample | Paraffin-embedded mouse spleen tissue; fixative not specified (datasheet PB9758) |
| Fixation | Image fixative and duration unreported (datasheet PB9758); 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 PB9758); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet PB9758) |
| Primary antibody | Rabbit anti-EIF4A2, 0.5-1μg/ml (datasheet PB9758) |
| Primary incubation | Overnight at 4 °C (datasheet PB9758) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet PB9758) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | EIF4A2-positive staining in adipocytes of adipose tissue (HPA tissue IHC: High). HPA tissue profile: Ubiquitous cytoplasmic expression. No signal in the no-primary control. |
💡Decision noteStart with EDTA pH 8.0 heat retrieval (datasheet: PB9758); the esophageal study also used EDTA pH 8.0 (PMC7471732 methods).
Section 2
What Is the Expected EIF4A2 Staining Pattern?
EIF4A2 should appear predominantly cytoplasmic in paraffin-section IHC, with staining across many cell types; HPA reports high staining in selected adipocytes, glandular cells, respiratory epithelium, and brain cell populations (HPA: ubiquitous cytoplasmic profile; HPA: listed High cells). HPA rates its tissue IHC profile Uncertain, pending external verification (HPA: reliability Uncertain). EIF4A2 has no transmembrane segment; UniProt does not annotate its subcellular location (UniProt Q14240: topology and subcellular record).
| Cytoplasmic staining in adipocytes or glandular cells, with visible cellular boundaries. | This fits the reported compartment and examples of High staining (HPA: ubiquitous cytoplasmic profile; HPA: adipocytes and glandular cells High). Score the stained cell population and intensity, then compare with a matched control; the HPA tissue profile remains Uncertain (HPA: reliability Uncertain). |
| Predominantly nuclear, membranous, or extracellular staining, with little cytoplasmic signal. | Treat this as a compartment mismatch requiring review, not proof of EIF4A2 localisation (HPA: ubiquitous cytoplasmic profile; HPA ICC-IF: cytosol Approved). UniProt supplies no subcellular annotation, so the mismatch alone cannot identify the source of the signal (UniProt Q14240: subcellular record). |
| Strong colour in an unexpected cell population while the expected cytoplasmic pattern is absent. | Check cross-reactivity and endogenous detection activity using controls (general IHC practice). HPA describes expression as ubiquitous and lists no negative tissue, so an unlisted cell type is not automatically a false positive (HPA: tissue profile; HPA: negative list empty). |
| Diffuse colour across tissue spaces or similar colour in cells and the surrounding section. | This is difficult to score as cellular EIF4A2 staining because the reported pattern is cytoplasmic (HPA: tissue profile). Review detection and background controls, including a primary-antibody omission control (general chromogenic IHC practice). |
| No cytoplasmic signal in a section containing an HPA-listed High cell population. | The result needs a run-quality check, but an HPA High listing is not a guarantee for every specimen (HPA: listed High cells; HPA: reliability Uncertain). Compare a positive-control section and detection controls before interpreting absence as biological loss (general IHC practice). |
💡Expected EIF4A2 appearanceCall a result consistent when discernible cytoplasmic staining occurs in the relevant cells, potentially strong in HPA-listed High populations; predominantly nuclear, membranous, or diffuse extracellular colour needs investigation (HPA: ubiquitous cytoplasmic profile; HPA: listed High cells; HPA: reliability Uncertain).
| Tissue and cell context | HPA reports low tissue RNA specificity and ubiquitous cytoplasmic IHC staining, with selected cell populations scored High (HPA: RNA specificity; HPA: tissue profile). Use those populations as comparators, not as a list of exclusive positives; HPA lists no negative tissue (HPA: negative list empty). |
| Antibody evidence | The tissue profile has medium agreement with RNA data and awaits external verification (HPA: reliability Uncertain). One listed antibody has IHC rated Uncertain; the other is listed as ICC Approved without an IHC rating (HPA: CAB011690; HPA: HPA068286). Interpret an isolated unusual pattern cautiously. |
| Protein topology and forms | EIF4A2 has no signal peptide, propeptide, or transmembrane segment; its annotated chain spans residues 1–407 (UniProt Q14240: processing and topology). Two isoforms and a phosphothreonine at residue 159 are recorded, but the supplied evidence does not show how either changes IHC staining (UniProt Q14240: isoforms and modified residues). |
| Fixation and retrieval | Target-specific fixation sensitivity and retrieval requirements are unreported in these payloads (UniProt Q14240; HPA: tissue and subcellular records). If a run fails, assess retrieval against the laboratory's established paraffin IHC controls (general IHC practice); do not infer an EIF4A2-specific fixation effect. |
| IF/ICC Q&A: what localisation is expected? | Cytosolic signal is the HPA-approved main location in ICC-IF (HPA: subcellular record). That annotation carries a caution because its supporting antibodies target proteins from multiple genes (HPA: subcellular caution). It supports a localisation comparison, not an IF/ICC protocol or IHC validation claim. |
| Situation | Likely cause | Next action |
|---|---|---|
| Cytoplasmic signal is absent in a positive-control section. | A failed staining run is possible; HPA's High populations supply comparators but its tissue profile is Uncertain (HPA: listed High cells; HPA: reliability Uncertain). | Check the control section, primary-antibody application, retrieval step, and detection reagents against the established paraffin IHC workflow (general IHC practice). Repeat the run before scoring the study section as negative. |
| Nuclear or membrane colour dominates. | The compartment disagrees with HPA's cytoplasmic tissue profile and approved cytosolic ICC-IF location (HPA: tissue profile; HPA: subcellular record). The source of the mismatch remains unresolved. | Review morphology and counterstain, then compare primary-antibody omission and positive controls (general IHC practice). Report the compartment observed rather than assigning it to EIF4A2 without further validation. |
| Unexpected cells stain strongly. | Cross-reactivity or endogenous detection activity is possible (general IHC practice). Because HPA calls expression ubiquitous, cell identity alone cannot establish a false positive (HPA: tissue profile). | Confirm which cells carry cytoplasmic signal; compare controls and a known HPA-listed High population in the same run (HPA: listed High cells; general IHC practice). |
| Colour is widespread without clear cell outlines. | Background or endogenous detection activity may obscure cellular staining (general chromogenic IHC practice); this cannot be confidently matched to HPA's cytoplasmic pattern (HPA: tissue profile). | Inspect a primary-antibody omission control and the detection-only background; adjust the established blocking or detection workflow as indicated by those controls (general IHC practice). |
| One tissue appears negative despite a reported High population. | HPA's High score describes its observed specimens and carries an Uncertain reliability rating; it does not establish positivity in every section (HPA: listed High cells; HPA: reliability Uncertain). | Verify that the named cell population is present and preserved, then compare a separate positive-control section from the same run (general IHC practice). Record absence without inferring EIF4A2 loss from this stain alone. |
| IHC and ICC-IF localisation seem inconsistent. | The methods and antibody evidence differ: tissue IHC is Uncertain, while the HPA ICC-IF cytosol assignment is Approved with a multiple-gene antibody caution (HPA: tissue reliability; HPA: subcellular record). | Compare the observed compartment and antibody validation status for each application (HPA: CAB011690 IHC Uncertain; HPA: HPA068286 ICC Approved). Assess each experiment with its own controls (general IHC/ICC practice). |
Experiment evidence
IHC & IF images showing EIF4A2 staining
PB9758 · IHCIHC analysis of eIF4A2 using anti-eIF4A2 antibody (PB9758). eIF4A2 was detected in a paraffin-embedded section of mouse spleen tissue. Heat
PB9758 · IHCIHC analysis of eIF4A2 using anti-eIF4A2 antibody (PB9758). eIF4A2 was detected in a paraffin-embedded section of rat intestine tissue. Heat
PB9758 · IFIF analysis of eIF4A2 using anti-eIF4A2 antibody (PB9758). eIF4A2 was detected in immunocytochemical section of A431 cell. Enzyme antigen re
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000156976-EIF4A2/tissue/cerebral+cortex#imid_3704064
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000156976-EIF4A2/tissue/colon#imid_3703990
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000156976-EIF4A2/tissue/liver#imid_3704007
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000156976-EIF4A2/tissue/kidney#imid_3704000
HPA · ICC-IF CACO-2Human Protein Atlas immunofluorescence, CACO-2, antibody HPA068286; green = EIF4A2, blue = nucleus, red = microtubules.
HPA · ICC-IF CACO-2Human Protein Atlas immunofluorescence, CACO-2, antibody HPA068286; green = EIF4A2, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for EIF4A2 IHC & IF
🧪Start with adipose tissue and score adipocytes, which are reported as High for EIF4A2 (HPA: High in adipocytes). HPA detects EIF4A2 in all 45 scored tissues, so there is no validated negative tissue; the no-primary and isotype controls provide the negative comparison, while any intact counterstained cells without DAB signal on the positive slide should remain free of chromogen (HPA: detected in all 45 scored tissues).
Positive control tissue: Adipose tissue (Adipocytes, HPA High)
Negative control tissue: None in HPA: EIF4A2 is detected in all 45 scored tissues. Use a no-primary (secondary-only) and an isotype control instead.
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show EIF4A2 in CACO-2, U-251MG, U2OS, with annotated localisation: Cytosol (approved) (HPA subcellular).
Technical controls: Run a secondary-only slide with the primary omitted, a concentration-matched rabbit IgG isotype control, and EIF4A2-knockout material as a biological specificity control (PB9758 caption: rabbit primary). Quench endogenous peroxidase for DAB detection and score adipocytes by their cellular rim rather than the empty lipid space (PB9758 caption: DAB detection; standard IHC practice).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected PB9758 paraffin-section caption does not state the fixative (PB9758 caption). That caption uses heat-mediated EDTA retrieval at pH 8.0, but does not establish that retrieval is required; the supplied evidence also does not show that frozen sections or IF/ICC are easier (PB9758 caption). Lipid extraction can leave empty spaces in adipocytes and complicate scoring (standard paraffin-section IHC practice).
HPA tissue IHC evidence for EIF4A2
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Uncertain — 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 |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | High | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | High | Protein (IHC) | HPA → |
| Appendix | Glandular cells | High | Protein (IHC) | HPA → |
| Breast | Glandular cells | High | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| None in HPA: EIF4A2 is detected in all 45 scored tissues. Use a no-primary (secondary-only) and an isotype control instead. | ||||
Section 3
Advanced EIF4A2 IHC Tips
Use the catalog antibody’s paraffin-section result as the IHC starting point, then assess cytoplasmic staining against controls and tissue morphology.
What retrieval should I try when EIF4A2 staining is weak in paraffin sections?
Start with heat-mediated retrieval in EDTA at pH 8.0 (datasheet PB9758). The selected mouse spleen image used this retrieval before incubation with 1 μg/ml primary antibody overnight at 4°C, so reproduce those documented conditions when evaluating weak staining (caption PB9758). Keep retrieval conditions and section handling consistent across the test and its controls, then compare cytoplasmic signal with tissue structure preserved (standard IHC practice; HPA: ubiquitous cytoplasmic expression). If staining remains weak, compare a small range of heating durations on matched sections before considering a different buffer or pH; the caption does not establish an alternative retrieval condition (standard IHC practice; caption PB9758).
Can I infer the best fixative for EIF4A2 from the published tissue staining?
Target-specific fixation sensitivity is unknown: the selected paraffin-section caption does not state a fixative, and the supplied tissue profile does not test fixation conditions (caption PB9758; HPA: tissue IHC profile). Record the fixative, fixation duration and processing history for every specimen before comparing staining intensity (standard IHC practice). If signal varies between batches, test matched material processed under controlled conditions while keeping EDTA at pH 8.0 and primary incubation unchanged (datasheet PB9758; standard IHC practice). Interpret any difference as a processing effect requiring validation, rather than attributing it to EIF4A2’s phosphorylation, isoforms or lack of a transmembrane segment (UniProt Q14240; standard IHC practice).
How should I assess nuclear or membranous staining for EIF4A2?
Expect predominantly cytoplasmic staining when reading chromogenic sections, while treating that expectation cautiously because the tissue IHC reliability is uncertain (HPA: ubiquitous cytoplasmic expression; HPA: uncertain reliability). A cytosolic location is approved in the subcellular record, but that record cautions that its antibodies target proteins from multiple genes (HPA: cytosol approved; HPA: subcellular caution). EIF4A2 has no transmembrane segment, so a crisp membrane-only pattern warrants investigation rather than immediate biological interpretation (UniProt Q14240 topology; standard IHC practice). Compare nuclear, membrane and cytoplasmic signal within intact cells, and review the same compartments in omission and detection controls before assigning a localisation (standard IHC practice).
Could an isoform or modified epitope explain inconsistent EIF4A2 IHC staining?
EIF4A2 has 2 recorded isoforms and a modified threonine at residue 159, but the supplied caption does not identify the antibody epitope or establish isoform-specific staining (UniProt Q14240; caption PB9758). Its helicase ATP-binding region spans residues 64–235 and its C-terminal region spans 246–407; these annotations alone cannot identify which region the antibody recognizes (UniProt Q14240 domains; caption PB9758). Check any available epitope documentation before interpreting staining differences as isoform expression or phosphorylation changes (standard IHC practice). For a disputed pattern, use an independent antibody with a documented, distinct epitope and appropriate tissue controls, then compare cellular distribution rather than staining intensity alone (standard IHC practice).
How can IF help check an ambiguous cytoplasmic IHC pattern?
Use IF as a complementary localisation check on separately prepared material, with a marker for the cell type being evaluated in the IHC section (standard IF practice). For example, a macrophage marker can test cell identity, while the expected EIF4A2 signal remains cytosolic and the tissue IHC profile carries uncertain reliability (standard IF practice; HPA: cytosol approved; HPA: uncertain reliability). Choose fluorophores and imaging channels after checking tissue autofluorescence, and include single-stain controls for multiplexing (standard IF practice). Because EIF4A2 lacks a transmembrane segment and the reported location is cytosolic, assess permeabilisation for access to intracellular epitopes; the supplied IHC caption does not establish an IF fixation or permeabilisation condition (UniProt Q14240 topology; HPA: cytosol approved; caption PB9758).
What should I check when DAB obscures EIF4A2 staining?
The selected paraffin-section method used 10% goat serum blocking, a biotinylated secondary antibody, a streptavidin–biotin complex and DAB detection (caption PB9758). If diffuse brown staining obscures cells, compare a primary-omission control with the test section and inspect whether deposit follows tissue edges, damaged areas or endogenous enzyme activity (standard IHC practice). Include a peroxidase block as part of the chromogenic workflow, and assess potential endogenous biotin interference when using the documented biotin-based detection system (standard IHC practice; caption PB9758). Adjust blocking, washing or detection only after identifying the likely source, then verify that interpretable cytoplasmic staining remains in intact cells (standard IHC practice; HPA: ubiquitous cytoplasmic expression).
How should I score EIF4A2 staining across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and viable tissue region before scoring, and use the same illumination, counterstain criteria and staining batch where possible (standard IHC practice). Record cytoplasmic intensity by category and the percentage of cells in each category; an H-score sums percentage multiplied by intensity and runs from 0–300 for a 0–3 scale (standard IHC practice; HPA: ubiquitous cytoplasmic expression). If tissue composition varies, report the percentage positive within the specified cell population or positive-cell density per mm² of viable tissue, normalised to the corresponding eligible cell count or area (standard IHC practice). Keep compartment scoring separate, and avoid treating staining intensity alone as a direct measure of protein abundance because the tissue IHC profile is uncertain (standard IHC practice; HPA: uncertain reliability).
When is an EIF4A2-positive IHC result convincing rather than artefactual?
A convincing result shows reproducible staining in intact cell cytoplasm, with morphology and control sections supporting the assignment (HPA: ubiquitous cytoplasmic expression; standard IHC practice). Investigate isolated membrane-only or strong nuclear-only staining because the approved subcellular location is cytosol and EIF4A2 lacks a transmembrane segment (HPA: cytosol approved; UniProt Q14240 topology). Exclude staining restricted to cut edges, necrotic regions or sites of endogenous peroxidase activity before calling cells positive (standard IHC practice). Because the tissue IHC assessment is uncertain and pending external verification, corroborate a surprising cell-type pattern independently before making a biological claim about EIF4A2 distribution (HPA: uncertain reliability; standard IHC practice).
Boster reagents
Best EIF4A2 / Eukaryotic initiation factor 4A-II IHC Antibodies
The catalog antibody has real IHC images from paraffin sections of mouse spleen, rat intestine, and human mammary cancer tissue, plus IF/ICC data from A431 cells (catalog image captions).
Real IHC data
Anti-eIF4A2 Antibody ®
Cat # PB9758
PB9758 is shown in paraffin sections of mouse spleen, rat intestine, and human mammary cancer tissue (PB9758 IHC image captions). It is also shown in A431 cells by IF/ICC (PB9758 IF image caption).
Which to pick: Choose PB9758 for paraffin-section IHC because its own images document staining in mouse, rat, and human tissue (PB9758 IHC image captions); the fixative is unreported (PB9758 IHC image captions). For IF/ICC, PB9758 is listed for both applications and has an A431 cell image (PB9758 catalog applications; PB9758 IF image caption). It is also the cross-species choice here because the catalog lists human, mouse, and rat reactivity; clonality is unreported (PB9758 catalog reactivity; PB9758 catalog clone field).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q14240 (IF4A2_HUMAN, Eukaryotic initiation factor 4A-II).
- Human Protein Atlas. EIF4A2 tissue IHC expression (reliability: Uncertain).
- Human Protein Atlas. EIF4A2 subcellular location (ICC-IF): Localized to the cytosol. Caution: Based on antibodies targeting proteins from multiple genes..
- Human Protein Atlas. EIF4A2 antibody validation summary (2 antibodies).
- High expression of eIF4A2 is associated with a poor prognosis in esophageal squamous cell carcinoma. Oncology letters 2020 — PMC7471732.
- Eukaryotic initiation factor 4A2 promotes experimental metastasis and oxaliplatin resistance in colorectal cancer. Journal of experimental & clinical cancer research : CR 2019 — PMC6518650.
- Long non-coding RNA LINC00520 promotes the proliferation and metastasis of malignant melanoma by inducing the miR-125b-5p/EIF5A2 axis. Journal of experimental & clinical cancer research : CR 2020 — PMC7254730.
- Increased mRNA translation delays tumour initiation and exposes a therapeutic vulnerability in lung cancer. Molecular cancer 2026 — PMC13366719.
- PubMed PMID:8521730 — UniProt-cited evidence.
- PubMed PMID:16641997 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for EIF4A2 (Eukaryotic initiation factor 4A-II). Expected localisation Ubiquitous cytoplasmic staining (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet PB9758). Validated IHC antibody: PB9758 at 0.5-1μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Adipose tissue (Adipocytes); negative None in HPA (detected in all 45 tissues); use no-primary + isotype controls. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded mouse spleen tissue; fixative not specified (datasheet PB9758); 2. Fixation: Image fixative and duration unreported (datasheet PB9758); 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 PB9758); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet PB9758); 8. Primary antibody: Rabbit anti-EIF4A2, 0.5-1μg/ml (datasheet PB9758); 9. Primary incubation: Overnight at 4 °C (datasheet PB9758). Watch for: Fixation: Keep fixation consistent across sections. (standard IHC practice; not target-specific); Caveat: Tissue staining has uncertain reliability; verify specificity (HPA tissue IHC); Isoform / epitope: Two isoforms; epitope coverage is unreported (UniProt; datasheet PB9758); Regulation: No specific expression regulator annotated (UniProt). Evidence: UniProt Q14240, HPA tissue IHC · Uncertain, HPA ICC-IF · cytosol. Caution: Based on antibodies targeting proteins from…, Datasheet PB9758, PMC7471732, PMC6518650. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/EIF4A2/immunohistochemistry-ihc-guide.