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CEBPE / CCAAT/enhancer-binding protein epsilon · IHC design guide
Design Immunohistochemistry for CEBPE
Plan chromogenic paraffin IHC for nuclear CEBPE staining in bone marrow hematopoietic cells (HPA tissue IHC). This guide covers controls and fixation, with starting IHC dilutions of 1:100–1:300 for A03884-1 or 1:50–1:200 for A03884-2 (datasheets).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
CEBPE 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 | Nucleus (UniProt) | |
| Staining pattern | Nuclear staining in bone marrow hematopoietic cells (HPA tissue IHC) | |
| Antigen retrieval | Tris-EDTA pH 9.0 HIER, 95–98 °C, 20 min (rule: nuclear antigen) | |
| Positive control | Bone marrow+1 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 | Endogenous peroxidase in marrow may mimic staining (standard IHC practice) | |
| Regulation | Myeloid stage-linked expression (UniProt) | |
| Isoform / epitope | No isoforms or cleavage reported; chain 1–281 (UniProt) |
Section 1
Recommended CEBPE IHC & IF Protocols
Compare the catalog antibody’s IHC-P protocol (datasheet) with 2 published CEBPE IHC methods (PMC12268938; PMC11375656).
| Sample | Paraffin-embedded human brain tissue; fixative not specified (datasheet A03884-1) |
| Fixation | Image fixative and duration unreported (datasheet A03884-1); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat-induced epitope retrieval in Tris-EDTA buffer, pH 9.0, 20 min at 95–98 °C (standard rule: nuclear antigen) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% normal serum of the secondary host, 30 min, room temperature (standard) |
| Primary antibody | Rabbit anti-CEBPE, 1:100-1:300 (datasheet A03884-1) |
| Primary incubation | Overnight at 4 °C (standard) |
| Detection | HRP-polymer secondary, DAB chromogen 5–10 min (standard) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | CEBPE-positive staining in hematopoietic cells of bone marrow (HPA tissue IHC: High). HPA tissue profile: Nuclear expression in bone marrow poietic cells. No signal in the no-primary control. |
💡Decision noteStart with Tris-EDTA pH 9.0 retrieval at 95–98 °C for 20 min (page retrieval rule; UniProt Q15744: nuclear localization).
Section 2
What Is the Expected CEBPE Staining Pattern?
CEBPE is a nuclear transcription factor with no transmembrane segment (UniProt Q15744). In paraffin-section IHC, expect nuclear staining chiefly in bone marrow hematopoietic cells, where HPA reports high staining and an Enhanced tissue pattern with high antibody–RNA consistency (HPA tissue IHC). Preleptotene spermatocytes in testis show medium staining (HPA tissue IHC). Interpret signal by both its compartment and the cells carrying it.
| Clear nuclear staining in bone marrow hematopoietic cells. | This matches the reported high bone marrow signal (HPA tissue IHC) and nuclear location (UniProt Q15744). Check that stained nuclei belong to hematopoietic cells; a positive-looking field alone does not establish the expected cell pattern. |
| Predominantly cytoplasmic or membrane staining in a bone marrow section. | This does not match the nuclear tissue-IHC profile (HPA tissue IHC; UniProt Q15744). Treat it as suspect staining and review controls and detection conditions before calling the section CEBPE-positive. Compartment mismatch alone does not identify the artefact's cause. |
| Strong staining in adipocytes or respiratory epithelial cells. | HPA reports CEBPE as not detected in adipocytes of adipose tissue and respiratory epithelial cells of bronchus (HPA tissue IHC). Signal in those specific cell types warrants checks for cross-reactivity or endogenous detection activity; do not extend this negative call to every cell in either tissue. |
| A diffuse chromogenic haze covers nuclei and surrounding tissue. | The reported pattern is nuclear in bone marrow hematopoietic cells (HPA tissue IHC). A haze that obscures cell boundaries cannot be scored as that pattern. General IHC practice is to compare a matched negative control and inspect blocking, washes, and detection background. |
| No nuclear signal in a bone marrow section expected to contain hematopoietic cells. | This conflicts with HPA's high staining in bone marrow hematopoietic cells (HPA tissue IHC). First verify that the relevant cells are present and identifiable, then assess assay controls. A blank section alone cannot distinguish missing target-bearing cells from assay failure. |
💡Expected CEBPE appearanceCall a convincing positive when bone marrow hematopoietic cells show clear, high nuclear staining (HPA tissue IHC; UniProt Q15744); dominant extranuclear staining or strong signal in HPA-negative cell types is suspect, not a CEBPE-positive pattern.
| Cell distribution | HPA reports high staining in bone marrow hematopoietic cells and medium staining in testis preleptotene spermatocytes (HPA tissue IHC). Select and score the relevant cells within each section; these levels do not imply uniform staining across either tissue. |
| Nuclear location | CEBPE is nuclear and lacks a transmembrane segment (UniProt Q15744). Nuclear location therefore anchors the IHC interpretation. UniProt topology does not establish an antigen retrieval setting or a fixation effect. |
| Strength of tissue-pattern evidence | HPA rates the tissue IHC profile Enhanced for antibody–RNA consistency, and lists two antibodies with Enhanced IHC validation (HPA tissue IHC; HPA antibodies). This supports the reported pattern; it does not validate every reagent or every unexpected signal. |
| Epitope and retrieval limits | The supplied sources give no catalog-antibody epitope, dilution, retrieval condition, or target-specific fixation sensitivity. Any retrieval adjustment is general paraffin-IHC assay optimization, not a demonstrated CEBPE-specific requirement. |
| Situation | Likely cause | Next action |
|---|---|---|
| Bone marrow nuclei are blank. | Relevant hematopoietic cells may be absent from the assessed area, or the assay may have failed; HPA reports high staining in those cells (HPA tissue IHC). | Confirm cell identity on the section and review the run's positive control. If the control also fails, optimize the general IHC workflow and use the catalog antibody's documented IHC-P conditions where available. |
| Signal is mainly cytoplasmic or at cell borders. | That distribution conflicts with nuclear CEBPE in tissue IHC (HPA tissue IHC; UniProt Q15744); its cause cannot be assigned from localisation alone. | Compare the same cell types with negative controls, inspect nonspecific detection, and rescore only clearly nuclear signal. Do not count extranuclear colour as confirmation. |
| Unexpected cells stain strongly. | For adipocytes in adipose tissue or respiratory epithelial cells in bronchus, HPA reports no detected staining (HPA tissue IHC). Cross-reactivity or endogenous detection activity is possible, not proven. | Verify the cell identity, compare negative controls, and check background from the chromogenic detection system before assigning target-specific staining. |
| Colour obscures nuclear boundaries throughout the section. | Diffuse background prevents assessment of the expected nuclear cell pattern (HPA tissue IHC); the source does not establish a CEBPE-specific background mechanism. | Use a matched negative control; review blocking, reagent concentration, washing, and detection time as general IHC troubleshooting steps. |
| Testis staining seems weaker than bone marrow. | Medium staining in preleptotene spermatocytes versus high staining in bone marrow hematopoietic cells is the reported pattern (HPA tissue IHC). | Compare the named cell types, not whole-section average colour. Investigate only if the expected cells lack interpretable nuclear signal or assay controls fail. |
| IF/ICC question: what localisation is expected? | HPA supports nucleoplasmic localisation; additional plasma-membrane and cytosolic assignments are uncertain (HPA subcellular ICC-IF). | Interpret a clear nucleoplasmic pattern as supported (HPA subcellular ICC-IF). Treat isolated membrane or cytosolic signal cautiously; it does not override the nuclear tissue-IHC expectation (HPA tissue IHC). |
Experiment evidence
IHC & IF images showing CEBPE staining
A03884-1 · IHCImmunohistochemistry analysis of paraffin-embedded human brain tissue, using C/EBP-epsilon Antibody. The picture on the right is blocked wit
A03884-1 · IFImmunofluorescence analysis of HeLa cells, using C/EBP-epsilon Antibody. The picture on the right is blocked with the synthesized peptide.
A03884-2 · IHCImmunohistochemistry (IHC) analyzes of C/EBP-ε (K70) pAb in paraffin-embedded human breast carcinoma tissue at 1:100.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000092067-CEBPE/tissue/cerebral+cortex#imid_992000
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000092067-CEBPE/tissue/colon#imid_991918
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000092067-CEBPE/tissue/liver#imid_991935
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000092067-CEBPE/tissue/kidney#imid_991928
HPA · ICC-IF JURKATHuman Protein Atlas immunofluorescence, JURKAT, antibody HPA002928; green = CEBPE, blue = nucleus, red = microtubules.
HPA · ICC-IF JURKATHuman Protein Atlas immunofluorescence, JURKAT, antibody HPA002928; green = CEBPE, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for CEBPE IHC & IF
🧪Run bone marrow first: hematopoietic cells should show nuclear staining (HPA: High in bone-marrow hematopoietic cells; UniProt Q15744: nucleus). Use adipose tissue as the negative tissue, scoring adipocytes, and expect cells outside the stained hematopoietic population on the bone-marrow slide to lack distinct nuclear signal (HPA: adipocytes Not detected in adipose tissue; HPA: bone-marrow hematopoietic cells High).
Positive control tissue: Bone marrow (Hematopoietic cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show CEBPE in JURKAT, REH, U2OS, A-431, U-251MG, with annotated localisation: Nucleoplasm (supported) (HPA subcellular).
Technical controls: Include no-primary (secondary-only) and host- and isotype-matched controls for a monoclonal antibody, or host-matched nonimmune IgG for a polyclonal antibody; use a CEBPE knockout specimen if available, or a peptide-block control as shown for the catalog antibody (standard IHC practice; A03884-1 tissue-IHC caption: peptide block). Block endogenous peroxidase in bone marrow before chromogenic detection and check background on the no-primary slide (standard IHC practice).
⚠️Feasibility: A target-specific fixation window and fixation effect are unreported in the supplied evidence; the A03884-1 paraffin-section caption does not report its fixative (A03884-1 tissue-IHC caption). Antigen-retrieval dependence is also unreported, so compare retrieval conditions using the positive and negative tissues (standard IHC practice; HPA: bone-marrow hematopoietic cells High, adipose-tissue adipocytes Not detected). The supplied evidence does not establish that frozen sections or IF are easier; bone-marrow endogenous peroxidase can complicate chromogenic scoring (standard IHC practice).
HPA tissue IHC evidence for CEBPE
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Enhanced — High 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 |
|---|---|---|---|---|
| Bone marrow | Hematopoietic cells | High | Protein (IHC) | HPA → |
| Testis | Preleptotene spermatocytes | Medium | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Appendix | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Breast | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced CEBPE IHC Tips
Use nuclear staining and the expected cell population to assess CEBPE in paraffin section IHC (UniProt Q15744; HPA tissue IHC).
How should I retrieve CEBPE in paraffin sections if nuclear staining is weak?
Start with Tris-EDTA pH 9.0 heat-induced retrieval at 95–98 °C for 20 minutes (page antigen-retrieval setting). Compare a known positive bone marrow section with the test section in the same run, scoring staining in hematopoietic-cell nuclei (HPA tissue IHC; standard IHC practice). If staining remains weak, vary heating time cautiously around the stated 20 minutes while checking tissue integrity and nonspecific staining (standard IHC practice). Keep antibody detection and chromogen development constant during that comparison so a retrieval change can be judged (standard IHC practice). The expected signal is nuclear, chiefly nucleoplasmic, which makes diffuse cytoplasmic color a poor endpoint for optimization (UniProt Q15744; HPA subcellular).
How should I troubleshoot variable CEBPE staining across differently fixed sections?
CEBPE-specific sensitivity to fixation is unknown from the supplied evidence; the selected antibody caption identifies paraffin-embedded human brain but does not state its fixative (catalog antibody A03884-1 caption). Record the fixative, fixation duration, section thickness, and storage history for each sample before comparing staining (standard IHC practice). Run sections with similar processing alongside a bone marrow positive control, where hematopoietic-cell nuclear staining is expected (HPA tissue IHC; standard IHC practice). If processing histories differ, compare morphology and nuclear signal after the stated Tris-EDTA pH 9.0 retrieval at 95–98 °C for 20 minutes (page antigen-retrieval setting; standard IHC practice). Avoid attributing a weak result to a CEBPE-specific fixation effect without a controlled comparison (standard IHC practice).
What staining compartment should I accept as CEBPE positive?
Score convincing nuclear staining as the primary CEBPE pattern: UniProt places the protein in the nucleus, and HPA supports a mainly nucleoplasmic location (UniProt Q15744; HPA subcellular). In bone marrow, examine hematopoietic-cell nuclei before treating color elsewhere as target signal (HPA tissue IHC). HPA also lists plasma membrane and cytosol signals as uncertain, while the record reports no transmembrane segment; do not use those compartments alone to call an IHC section positive (HPA subcellular; UniProt Q15744 topology). Check an adjacent negative-control section for comparable extranuclear color and review nuclear morphology with a counterstain (standard IHC practice). Report any reproducible extranuclear pattern separately from the nuclear score (standard IHC practice).
Could epitope placement or modification explain discordant CEBPE staining?
The supplied record lists 0 isoforms, a bZIP domain at residues 204–267, and phosphoserine at residue 181 (UniProt Q15744). The catalog antibody's epitope is not mapped in this payload, so those features cannot establish whether retrieval or modification changes antibody binding (catalog antibody A03884-1 caption; UniProt Q15744). If results disagree between antibodies, compare their documented immunogens or mapped epitopes before interpreting the difference biologically (standard IHC practice). Keep tissue processing and the page's Tris-EDTA pH 9.0, 95–98 °C, 20-minute retrieval consistent during that comparison (page antigen-retrieval setting; standard IHC practice). Use nuclear staining in expected cells and appropriate controls to assess each antibody independently (HPA tissue IHC; standard IHC practice).
How can I use IF to check a CEBPE IHC pattern?
Use IF as a separate assay to examine whether CEBPE signal falls within nuclei of the expected cells (UniProt Q15744; HPA subcellular; standard IF practice). Multiplex a nuclear stain with a validated hematopoietic-cell marker when assessing bone marrow, since HPA reports high expression in those cells (HPA tissue IHC; standard IF practice). Choose a far-red fluorophore after checking the tissue's autofluorescence and include single-stain controls for channel bleed-through (standard IF practice). Permeabilize fixed cells or sections to let antibody reach the nucleoplasmic target, for example with 0.1% Triton X-100 for 5 minutes, then verify that morphology remains intact (HPA subcellular; standard IF practice). IF preparation and antibody performance require their own controls and optimization (standard IF practice).
How do I distinguish nonspecific chromogen from CEBPE staining?
First compare the suspected signal with the expected nuclear pattern in hematopoietic cells and with a negative-control section processed through detection (HPA tissue IHC; standard IHC practice). Block endogenous peroxidase before horseradish-peroxidase detection, for example with 3% hydrogen peroxide for 10 minutes, following the detection system's instructions (standard chromogenic IHC practice). If diffuse color persists, assess primary-antibody concentration, blocking, washing, and DAB development time one variable at a time (standard IHC practice). The selected catalog image includes a synthesized-peptide-blocked view of paraffin-embedded human brain; that comparison is useful context but does not by itself establish staining specificity in every tissue (catalog antibody A03884-1 caption; standard IHC interpretation).
How should I quantify CEBPE across bone marrow sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the hematopoietic-cell population and score its nuclei, because HPA reports high CEBPE staining in bone marrow hematopoietic cells and describes nuclear expression there (HPA tissue IHC). Report the percentage of positive nuclei and, if intensity is reproducible, an H-score from 0–300 using intensity categories 0–3 (standard IHC scoring practice). Normalize positive counts to the number of evaluable cells in that population, or report positive-cell density per mm² of evaluable tissue (standard IHC quantification practice). Keep threshold, counterstain, imaging, and sampling rules fixed across cases, and exclude folds, necrosis, and section edges from scored regions (standard IHC practice).
When should I doubt an apparent CEBPE-positive IHC result?
A plausible positive shows nuclear staining in an appropriate cell population: HPA reports high staining in bone marrow hematopoietic cells and supported nucleoplasmic localization (HPA tissue IHC; HPA subcellular). Question isolated cytoplasmic or membrane color because HPA labels those additional locations uncertain and UniProt reports no transmembrane segment (HPA subcellular; UniProt Q15744 topology). Inspect section edges and necrotic regions for staining artifacts, and use a negative-control section to evaluate detection background or endogenous enzyme activity (standard IHC practice). The catalog antibody's peptide-blocked brain image provides a control for that image, while HPA reports no detectable staining in the listed cerebral-cortex endothelial cells; identify the stained cell population before generalizing either observation (catalog antibody A03884-1 caption; HPA tissue IHC).
Boster reagents
Best CEBPE / CCAAT/enhancer-binding protein epsilon IHC Antibodies
Two anti-CEBPE antibodies list human, mouse and rat reactivity and show paraffin-section IHC images; A03884-1 also shows IF staining in HeLa cells (catalog reactivity; catalog image captions).
Real IHC data
Anti-C/EBP epsilon CEBPE Antibody
Cat # A03884-1
Real IHC data
Anti-C/EBP-Epsilon (K70) CEBPE Antibody
Cat # A03884-2
A03884-1 has IHC images of paraffin-embedded human brain tissue and an IF image of HeLa cells, each with peptide blocking (catalog image captions). A03884-2 has an IHC image of paraffin-embedded human breast carcinoma at 1:100 and lists IF as an application (catalog IHC caption; catalog applications).
Which to pick: For tissue IHC, choose A03884-1 for its peptide-blocked human brain image or A03884-2 for its human breast carcinoma image; both captions specify paraffin-embedded sections, but neither reports the fixative (catalog IHC captions). For IF/ICC, A03884-1 has a peptide-blocked HeLa IF image, while A03884-2 lists IF without an IF image (catalog IF captions; catalog applications). Both list human, mouse and rat reactivity; A03884-1 is listed as polyclonal and A03884-2 is described as pAb, so confirm staining in the intended species and tissue (catalog reactivity; catalog dilution_raw; A03884-2 IHC caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q15744 (CEBPE_HUMAN, CCAAT/enhancer-binding protein epsilon).
- Human Protein Atlas. CEBPE tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. CEBPE subcellular location (ICC-IF): Mainly localized to the nucleoplasm. In addition localized to the plasma membrane and cytosol..
- Human Protein Atlas. CEBPE antibody validation summary (2 antibodies).
- Deliver CEBPE via cartilage targeting Lipid nanoparticle to block CEBPE-LTF-STAT3 positive feedback loop for efficient treatment of cartilage endplate degeneration. Materials today. Bio 2025 — PMC12268938.
- Comprehensive Analysis of CCAAT/Enhancer Binding Protein Family in Ovarian Cancer. Cancer informatics 2024 — PMC11375656.
- PubMed PMID:8661101 — UniProt-cited evidence.
- PubMed PMID:9032264 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for CEBPE (CCAAT/enhancer-binding protein epsilon). Expected localisation Nucleus (UniProt). Antigen retrieval: Tris-EDTA pH 9.0 HIER, 95–98 °C, 20 min (rule: nuclear antigen). Validated IHC antibody: A03884-1 at 1:100-1:300 (the product's own IHC validation image is shown). Controls (HPA): positive Bone marrow (Hematopoietic cells); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human brain tissue; fixative not specified (datasheet A03884-1); 2. Fixation: Image fixative and duration unreported (datasheet A03884-1); 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-induced epitope retrieval in Tris-EDTA buffer, pH 9.0 , 20 min at 95–98 °C…; 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% normal serum of the secondary host, 30 min , room temperature (standard); 8. Primary antibody: Rabbit anti-CEBPE, 1:100-1:300 (datasheet A03884-1); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across paraffin sections. (standard IHC practice; not target-specific); Caveat: Endogenous peroxidase in marrow may mimic staining (standard IHC practice); Isoform / epitope: No isoforms or cleavage reported; chain 1–281 (UniProt); Regulation: Myeloid stage-linked expression (UniProt). Evidence: UniProt Q15744, HPA tissue IHC · Enhanced, HPA ICC-IF · Mainly localized to the nucleoplasm. In addition localized to the…, Datasheet A03884-1, PMC12268938, PMC11375656. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/CEBPE/immunohistochemistry-ihc-guide.