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EIF4B / Eukaryotic translation initiation factor 4B · IHC design guide
Design Immunohistochemistry for EIF4B
Plan chromogenic EIF4B IHC in paraffin sections with the IHC-validated antibody at 2–5 μg/ml (datasheet A00846-1). Expect general cytoplasmic staining, with high signal reported in cervical squamous epithelial cells and duodenal glandular cells (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
EIF4B 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 cytoplasmic staining (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic staining in squamous and glandular cells (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A00846-1) | |
| Positive control | Cervix+4 more · see all | |
| Negative control | Adipose tissue+3 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent across sections (standard IHC practice; not target-specific) | |
| Caveat | Staining and RNA show medium consistency (HPA tissue IHC) | |
| Regulation | Low tissue specificity (HPA tissue RNA) | |
| Isoform / epitope | 2 isoforms; verify epitope coverage (UniProt) |
Section 1
Recommended EIF4B IHC & IF Protocols
The catalog antibody protocol (datasheet A00846-1) is accompanied by published IHC methods for human brain sections (PMC8339060) and tissue microarrays (PMC5827760).
| Sample | Paraffin-embedded human cervical cancer tissue; fixative not specified (datasheet A00846-1) |
| Fixation | Image fixative and duration unreported (datasheet A00846-1); 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 A00846-1); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A00846-1) |
| Primary antibody | Rabbit anti-EIF4B, 2-5 μg/ml (datasheet A00846-1) |
| Primary incubation | Overnight at 4 °C (datasheet A00846-1) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet A00846-1) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | EIF4B-positive staining in squamous epithelial cells of cervix (HPA tissue IHC: High). HPA tissue profile: General cytoplasmic expression. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA retrieval at pH 8.0 (datasheet A00846-1); neither published excerpt specifies retrieval conditions (PMC8339060; PMC5827760).
Section 2
What Is the Expected EIF4B Staining Pattern?
EIF4B should show predominantly cytoplasmic staining in paraffin tissue sections, especially in squamous epithelial cells of cervix, esophagus, oral mucosa and vagina, and glandular cells of duodenum, rectum and small intestine (HPA tissue IHC). HPA rates its tissue staining Approved, with medium consistency against RNA data and external verification pending (HPA tissue IHC). EIF4B has no transmembrane segment; UniProt does not annotate a subcellular location (UniProt P23588 topology; UniProt P23588 subcellular).
| Strong cytoplasmic staining in squamous epithelial cells of cervix, esophagus, oral mucosa or vagina (HPA tissue IHC). | This matches the cell type, compartment and High staining reported in those tissues (HPA tissue IHC). Assess the epithelial cells themselves; staining elsewhere on the section does not substitute for the expected pattern (HPA tissue IHC; general IHC practice). |
| Predominantly nuclear staining in cells expected to be positive (HPA tissue IHC). | A nuclear-dominant pattern conflicts with HPA's general cytoplasmic tissue profile and should prompt an artefact check (HPA tissue IHC; general IHC practice). UniProt provides no subcellular annotation that would independently support a nuclear interpretation (UniProt P23588 subcellular). |
| Strong staining confined to adipocytes, cholangiocytes, smooth muscle cells or chondrocytes (HPA tissue IHC). | HPA reports these specified cells as Not detected in adipose tissue, liver, smooth muscle and soft tissue, respectively (HPA tissue IHC). Check for cross-reactivity or endogenous detection activity before calling their signal EIF4B; these observations do not make every cell in those tissues a negative control (HPA tissue IHC; general IHC practice). |
| Diffuse color across cells and surrounding tissue, without a clear cytoplasmic boundary (general IHC practice). | This is difficult to score as specific staining against HPA's general cytoplasmic profile (HPA tissue IHC). Background may arise from nonspecific antibody binding or the detection system; inspect a matched control and the counterstained morphology (general IHC practice). |
| No convincing signal in squamous epithelium of esophagus or glandular cells of duodenum (HPA tissue IHC). | HPA reports High staining in those cells, so the result warrants a technical review (HPA tissue IHC; general IHC practice). It does not by itself prove EIF4B absence: review tissue identity, controls and the IHC workflow before interpreting a negative result (general IHC practice). |
💡Expected EIF4B appearanceCall positive a clear, mainly cytoplasmic signal in the expected squamous or glandular cells, with High staining in HPA-listed high-expression examples; nuclear-dominant or cell-free diffuse color is suspect (HPA tissue IHC; general IHC practice).
| Tissue and cell choice | HPA reports High staining in selected squamous epithelia and intestinal glandular cells, but Low staining in several neuronal and glandular cell groups (HPA tissue IHC). Choose the exact cell population when judging a control; a weak result in a Low group is less informative than one in a High group (HPA tissue IHC; general IHC practice). |
| Evidence and antibody validation | The HPA tissue profile is Approved, with medium consistency against RNA expression and external verification pending (HPA tissue IHC). HPA046164 is IHC Approved; its ICC Enhanced status applies to ICC, while HPA039072 has ICC Approved and no listed IHC status (HPA antibodies). Do not transfer ICC validation to paraffin IHC (HPA antibodies). |
| Isoforms and epitope coverage | UniProt lists two EIF4B isoforms and an RRM at residues 96–173, but the supplied record gives no antibody epitope (UniProt P23588 isoforms; UniProt P23588 domains). Whether an antibody recognizes both isoforms cannot be concluded from these facts; use the actual antibody documentation when interpreting discordant staining (UniProt P23588 isoforms; general IHC practice). |
| Phosphorylation and processing | UniProt lists multiple modified residues, including phosphoserines at 93, 192, 207, 219 and 283; it lists one 1–611 chain and no signal peptide or propeptide (UniProt P23588 modified residues; UniProt P23588 processing). These annotations do not establish a phospho-dependent staining pattern or target-specific fixation sensitivity (UniProt P23588; general IHC practice). |
| IF/ICC Q: What localisation is expected on its separate guide page? | A: HPA places EIF4B mainly in endoplasmic reticulum and cytosol, with additional plasma membrane localisation; those locations are Approved in ICC-IF (HPA subcellular). This cell-imaging evidence can inform interpretation, but it does not turn a paraffin IHC result into an IF protocol or establish identical visual detail in tissue sections (HPA subcellular; general IHC practice). |
| Situation | Likely cause | Next action |
|---|---|---|
| Expected epithelium or intestinal glands show no signal (HPA tissue IHC). | The High HPA pattern was not reproduced; tissue selection or an IHC workflow step may need review (HPA tissue IHC; general IHC practice). | Confirm the expected cells on the counterstain, review a known-positive section and check the antibody and detection workflow (HPA tissue IHC; general IHC practice). |
| Nuclei dominate while cytoplasm is faint (HPA tissue IHC). | This differs from the general cytoplasmic tissue pattern; UniProt supplies no independent nuclear localisation annotation (HPA tissue IHC; UniProt P23588 subcellular). | Compare the signal with morphology and controls, then review staining and scoring before assigning EIF4B localisation (general IHC practice). |
| Color appears mainly in HPA-listed Not detected cell groups (HPA tissue IHC). | Cross-reactivity or endogenous detection activity is possible; the HPA negatives refer to specified cells, not whole tissues (HPA tissue IHC; general IHC practice). | Check the cell identity and compare appropriate negative and detection controls with an expected positive cell population (HPA tissue IHC; general IHC practice). |
| Diffuse haze obscures cell boundaries (general IHC practice). | Nonspecific binding or detection background can prevent reliable cytoplasmic scoring (general IHC practice). | Inspect matched controls and review blocking, antibody dilution, washes and detection conditions as general IHC workflow checks; no EIF4B-specific settings are supplied (general IHC practice). |
| Only weak staining appears in a chosen tissue (HPA tissue IHC). | HPA reports Low staining in some cell groups, including colon glandular cells, so the chosen cells may offer limited contrast (HPA tissue IHC). | Compare the exact cell type with its HPA level and assess a High-staining reference such as duodenal glandular cells before judging assay performance (HPA tissue IHC; general IHC practice). |
| A change in antigen retrieval seems to change staining (general IHC practice). | Target-specific fixation effects are not established by the supplied assay evidence. Verify with a matched IHC source before attributing a result to fixation. | Compare conditions with the same control tissue and score cell type, compartment and background together; avoid attributing a change to a specific EIF4B epitope without antibody evidence (HPA tissue IHC; general IHC practice). |
Experiment evidence
IHC & IF images showing EIF4B staining
A00846-1 · IHCIHC analysis of EIF4B using anti-EIF4B antibody (A00846-1). EIF4B was detected in a paraffin-embedded section of human cervical cancer tissu
A00846-1 · IHCIHC analysis of EIF4B using anti-EIF4B antibody (A00846-1). EIF4B was detected in a paraffin-embedded section of human colorectal cancer tis
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000063046-EIF4B/tissue/cerebral+cortex#imid_15104722
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000063046-EIF4B/tissue/colon#imid_15104656
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000063046-EIF4B/tissue/liver#imid_15104649
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000063046-EIF4B/tissue/kidney#imid_15104650
HPA · ICC-IF HeLaHuman Protein Atlas immunofluorescence, HeLa, antibody HPA039072; green = EIF4B, blue = nucleus, red = microtubules.
HPA · ICC-IF HeLaHuman Protein Atlas immunofluorescence, HeLa, antibody HPA039072; green = EIF4B, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for EIF4B IHC & IF
🧪Run cervix first: squamous epithelial cells should stain strongly (HPA: High in cervical squamous epithelial cells). Run adipose tissue as the biological low-expression comparison (HPA: Not detected in adipocytes); within the cervix section, cells used as an internal negative reference should show counterstain without specific DAB signal, but no cervical cell type is identified as negative in the supplied HPA rows.
Positive control tissue: Cervix (Squamous epithelial cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show EIF4B in HeLa, MCF-7, U2OS, HEK293, RPTEC/TERT1, hTERT-RPE1 (serum starved), U2OS, siRNA 2 (10x), U2OS, scrambled (10x), with annotated localisation: Endoplasmic reticulum (approved), Cytosol (approved) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control; a nonimmune rabbit IgG control matched to the primary antibody’s class and clonality where known; and EIF4B knockout cells processed as a paraffin cell block or a validated peptide-block control (standard IHC practice; product caption: rabbit anti-EIF4B). Block endogenous peroxidase and check endogenous biotin background because the reported cervical-cancer assay used a biotin-based detection system with DAB (product caption: biotinylated secondary, SABC and DAB).
⚠️Feasibility: The selected A00846-1 paraffin-section caption does not state a fixative, and no target-specific fixation window or fixation effect is reported in the supplied evidence (product caption: fixative unreported). Heat retrieval in EDTA at pH 8.0 was used for that IHC example, but retrieval dependence has not been established; frozen-section performance is unreported, while HPA provides ICC-IF images that require separate IF optimisation (product caption: EDTA retrieval; HPA: ICC-IF images). No cervix-specific artefact is documented in the supplied evidence; assess background from the biotin-based DAB detection on the actual section (product caption: SABC and DAB; standard IHC practice).
HPA tissue IHC evidence for EIF4B
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 |
|---|---|---|---|---|
| Cervix | Squamous epithelial cells | High | Protein (IHC) | HPA → |
| Duodenum | Glandular cells | High | Protein (IHC) | HPA → |
| Esophagus | Squamous epithelial cells | High | Protein (IHC) | HPA → |
| Oral mucosa | Squamous epithelial cells | High | Protein (IHC) | HPA → |
| Rectum | 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 → |
| Liver | Cholangiocytes | Not detected | Protein (IHC) | HPA → |
| Smooth muscle | Smooth muscle cells | Not detected | Protein (IHC) | HPA → |
| Soft tissue | Chondrocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced EIF4B IHC Tips
Troubleshoot EIF4B staining in paraffin-section chromogenic IHC using the catalog antibody’s documented workflow and the reported tissue patterns (caption A00846-1; HPA tissue IHC).
Which retrieval condition should I try first for weak EIF4B staining?
Start with heat-mediated retrieval in EDTA at pH 8.0 for paraffin sections (datasheet A00846-1). If staining remains weak, compare an alternative retrieval condition on adjacent sections while keeping antibody concentration, detection, and development constant (standard IHC practice). The pictured workflow used 2 μg/ml primary antibody overnight at 4°C, followed by biotinylated secondary antibody for 30 minutes at 37°C and DAB development (caption A00846-1). Include a no-primary control and compare cytoplasmic signal in squamous epithelial cells with the expected pattern before interpreting stronger staining as improved retrieval (HPA: general cytoplasmic expression; HPA: High in cervix squamous epithelial cells; standard IHC practice).
Could fixation explain inconsistent EIF4B staining between paraffin sections?
For paraffin-section IHC, target-specific sensitivity of EIF4B to fixation is unknown; the selected image states that the section is paraffin embedded but does not report a fixative (caption A00846-1). Record fixation method and duration for every specimen, and compare matched sections processed through the same EDTA pH 8.0 retrieval and 2 μg/ml primary incubation before attributing differences to biology (datasheet A00846-1; caption A00846-1; standard IHC practice). Keep section thickness and DAB development consistent across the comparison (standard IHC practice). A weak or absent stain after a protocol change warrants a controlled repeat with positive and no-primary controls before attributing it to fixation (standard IHC practice).
How should I assess EIF4B staining that appears restricted to the cell membrane?
Expect predominantly cytoplasmic staining in tissue sections, while assessing apparent membrane-associated signal carefully (HPA: general cytoplasmic expression; HPA subcellular: approved ER, cytosol and additional plasma membrane). EIF4B has no transmembrane segment, so a crisp membrane-only outline warrants scrutiny before it serves as a positive criterion (UniProt P23588 topology; standard IHC practice). Compare the signal with high staining in cervical squamous epithelium and absent staining in adipocytes under matched EDTA pH 8.0 retrieval and DAB settings (HPA: High in cervix squamous epithelial cells; HPA: Not detected in adipocytes; datasheet A00846-1; standard IHC practice). Check nuclear counterstain and cell borders before assigning a compartment (standard IHC practice).
Can this stain distinguish EIF4B isoforms or phosphorylation states?
First establish which EIF4B sequence the antibody recognizes; the selected caption supplies the SKU but no epitope map (caption A00846-1). EIF4B has 2 isoforms, an RNA recognition motif at residues 96–173, and multiple modified residues including phosphoserines at 93 and 192 (UniProt P23588). Without mapped epitope and isoform coverage, a staining change cannot be assigned confidently to splice form or phosphorylation state (UniProt P23588; standard IHC interpretation). Compare matched sections under the same EDTA pH 8.0 retrieval and 2 μg/ml antibody conditions, then seek an independent epitope or orthogonal assay if the biological claim depends on isoform specificity (datasheet A00846-1; caption A00846-1; standard IHC practice).
How can IF help assess an EIF4B IHC localisation result?
Use IF/ICC as a separate validation readout for the IHC finding: multiplex EIF4B with a cell-type marker such as cytokeratin when examining squamous epithelium (HPA: High in cervix squamous epithelial cells; standard IF practice). Choose a far-red fluorophore when tissue autofluorescence overlaps shorter-wavelength channels, and include single-stain and no-primary controls (standard IF practice). Permeabilise for access to the reported cytosolic and endoplasmic-reticulum signal; the antibody’s epitope and its side of any membrane are unreported (HPA subcellular: approved cytosol and ER; caption A00846-1; standard IF practice). Keep the IHC comparison anchored to EDTA pH 8.0 retrieval and the recorded DAB workflow (datasheet A00846-1; caption A00846-1).
What should I check when EIF4B DAB staining looks diffuse?
If DAB appears diffuse, run no-primary and secondary-only controls to assess nonspecific detection, then inspect tissue away from cut edges and damaged areas (standard IHC practice). The pictured method uses 10% goat serum blocking, 2 μg/ml rabbit primary overnight at 4°C, biotinylated secondary for 30 minutes at 37°C, and streptavidin-biotin detection with DAB (caption A00846-1). Apply an endogenous peroxidase block and consider endogenous biotin controls where relevant to this detection system (standard IHC practice). Compare cellular cytoplasmic signal with negative areas, including adipocytes, before adjusting antibody concentration or DAB development time (HPA: general cytoplasmic expression; HPA: Not detected in adipocytes; standard IHC practice).
How should I quantify EIF4B across sections with different cell composition? ⚠ ANSWER MARKED FOR VERIFICATION
Score EIF4B in a predefined cell population and cytoplasmic compartment, using an H-score from 0–300 based on percentages of cells at intensity 0–3 (HPA: general cytoplasmic expression; standard IHC scoring). Report percentage positive cells alongside the score, with the positivity threshold set from a no-primary control before reading study sections (standard IHC practice). Normalise to the number of viable target cells or tissue area, and keep EDTA pH 8.0 retrieval, primary exposure, and DAB development matched across slides (datasheet A00846-1; standard IHC practice). Record epithelial and stromal compartments separately so changes in tissue composition do not masquerade as changes in EIF4B expression (standard IHC practice).
Which findings support a true EIF4B positive rather than an artefact?
Call staining credible when it is cellular and mainly cytoplasmic, occurs in the intended cell population, and exceeds matched no-primary background (HPA: general cytoplasmic expression; standard IHC practice). Use cervical squamous epithelial cells as a high-staining comparator and adipocytes as a not-detected comparator, while evaluating specificity with controls (HPA: High in cervix squamous epithelial cells; HPA: Not detected in adipocytes; standard IHC practice). Discount edge staining, necrotic deposits, and DAB signal persisting in the no-primary control; check endogenous peroxidase and biotin contributions to the pictured detection scheme (caption A00846-1; standard IHC practice). Interpret intensity differences cautiously because HPA reports medium RNA concordance and pending external verification (HPA tissue IHC).
Boster reagents
Best EIF4B / Eukaryotic translation initiation factor 4B IHC Antibodies
The catalog includes antibodies reactive with human, mouse and rat (catalog: reactivity). IHC images show EIF4B in human paraffin sections; no IF images are supplied (catalog: image captions).
Real IHC data
Anti-EIF4B Antibody ®
Cat # A00846-1
A00846-1 has IHC images from human cervical cancer, colorectal cancer, thyroid papillary carcinoma and tonsil paraffin sections (A00846-1: IHC image captions). P00846 is listed for IHC and recognizes phospho-eIF4B S406, but has no IHC image supplied (P00846: applications; title; image captions).
Which to pick: For tissue IHC, choose A00846-1 when an illustrated paraffin-section protocol matters: its caption specifies 2 μg/ml and EDTA retrieval at pH 8.0; the fixative is unreported (A00846-1: IHC image caption). Choose P00846 when the question is specifically S406 phosphorylation; it is a rabbit monoclonal with a listed IHC dilution of 1:50–1:200, but no IHC image is supplied (P00846: title; host; clone; dilution; image captions). Neither SKU lists IF/ICC validation; both list human, mouse and rat reactivity, while A00846-1’s pictured IHC samples are human only (catalog: applications; reactivity; A00846-1 IHC image captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P23588 (IF4B_HUMAN, Eukaryotic translation initiation factor 4B).
- Human Protein Atlas. EIF4B tissue IHC expression (reliability: Approved).
- Human Protein Atlas. EIF4B subcellular location (ICC-IF): Mainly localized to the endoplasmic reticulum and cytosol. In addition localized to the plasma membrane..
- Human Protein Atlas. EIF4B antibody validation summary (2 antibodies).
- Casein Kinase 2 dependent phosphorylation of eIF4B regulates BACE1 expression in Alzheimer's disease. Cell death & disease 2021 — PMC8339060.
- EIF4B Ser93 phosphorylation by ERK2 promotes epithelial-mesenchymal transition to drive colorectal cancer metastasis. Cell death & disease 2026 — PMC12877161.
- Loss of SLC25A11 causes suppression of NSCLC and melanoma tumor formation. EBioMedicine 2019 — PMC6413681.
- Fatty Acid Synthase induced S6Kinase facilitates USP11-eIF4B complex formation for sustained oncogenic translation in DLBCL. Nature communications 2018 — PMC5827760.
- PubMed PMID:2390971 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
- PubMed PMID:16541075 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for EIF4B (Eukaryotic translation initiation factor 4B). Expected localisation General cytoplasmic staining (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A00846-1). Validated IHC antibody: A00846-1 at 2-5 μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Cervix (Squamous epithelial cells); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human cervical cancer tissue; fixative not specified (datasheet A00846-1); 2. Fixation: Image fixative and duration unreported (datasheet A00846-1); 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 A00846-1); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A00846-1); 8. Primary antibody: Rabbit anti-EIF4B, 2-5 μg/ml (datasheet A00846-1); 9. Primary incubation: Overnight at 4 °C (datasheet A00846-1). Watch for: Fixation: Keep fixation consistent across sections (standard IHC practice; not target-specific); Caveat: Staining and RNA show medium consistency (HPA tissue IHC); Isoform / epitope: 2 isoforms; verify epitope coverage (UniProt); Regulation: Low tissue specificity (HPA tissue RNA). Evidence: UniProt P23588, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the endoplasmic reticulum and cytosol. In…, Datasheet A00846-1, PMC8339060, PMC5827760. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/EIF4B/immunohistochemistry-ihc-guide.