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E2F1 / Transcription factor E2F1 · IHC design guide
Design Immunohistochemistry for E2F1
Plan paraffin-section E2F1 IHC around nuclear staining in most tissues (HPA tissue IHC). Bone marrow hematopoietic cells offer a high-staining reference, while glomerular cells have no detected signal (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
E2F1 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 | Nuclear staining in most tissues (HPA tissue IHC) | |
| Staining pattern | Nuclear signal 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 | Adrenal gland+4 more · see all | |
| Negative control | Kidney+2 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); Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image M00257-1) | |
| Caveat | Glomerular cells have no detected signal (HPA tissue IHC) | |
| Regulation | G1–S activity; abundance regulation unreported (UniProt) | |
| Isoform / epitope | 0 annotated isoforms; no processing (UniProt) |
Section 1
Recommended E2F1 IHC & IF Protocols
The catalog antibody’s IHC-P protocol is accompanied by published E2F1 staining methods for consecutive tissue sections and harvested tumor specimens (PMC5362443; PMC11136682).
| Sample | Tissue sections; selected-image fixative not specified (standard IHC workflow) |
| Fixation | Image fixative and duration unreported (datasheet M00257-1); 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-E2F1, 1:200-1:1000 (datasheet M00257-1) |
| 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 | E2F1-positive staining in glandular cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: Nuclear expression in most tissues. No signal in the no-primary control. |
💡Decision noteStart with Tris-EDTA pH 9.0 retrieval at 95–98 °C for 20 min for nuclear E2F1 (page retrieval rule; UniProt: nucleus).
Section 2
What Is the Expected E2F1 Staining Pattern?
Nuclear, non-TM E2F1 (UniProt Q01094) stains marrow hematopoietic cells (HPA: High; Enhanced IHC).
| Nuclear stain in marrow hematopoietic cells (HPA: High; nuclear profile) | Expected positive pattern (HPA: High in marrow hematopoietic cells). |
| Predominantly cytoplasmic stain (HPA: nuclear profile) | Unexpected location; assess nonspecific staining (UniProt Q01094: nucleus). |
| Stain in skeletal muscle myocytes (HPA: Not detected) | Unexpected; consider cross-reactivity or endogenous enzyme activity (general IHC practice). |
| Diffuse stain across the section (general IHC practice) | Likely background; review blocking and washes (general IHC practice). |
| No stain in marrow hematopoietic cells (HPA: High) | Possible false negative; check controls and retrieval (general IHC practice). |
💡Expected E2F1 appearanceStrong nuclear marrow hematopoietic stain (HPA: High); cytoplasmic stain is suspect (UniProt Q01094).
| Tissue contrast (HPA tissue IHC) | Marrow hematopoietic cells: High; skeletal myocytes: Not detected (HPA tissue IHC). |
| Subcellular location (UniProt Q01094) | Score nuclear signal; E2F1 has no transmembrane segment (UniProt Q01094). |
| IF/ICC: where is E2F1? (HPA ICC-IF) | Mainly nucleoplasm (supported); centrosome also reported (uncertain) (HPA ICC-IF). |
| Antibody validation (HPA antibodies) | CAB000329: IHC Enhanced; HPA008003 and CAB019308: IHC Approved (HPA antibodies). |
| Situation | Likely cause | Next action |
|---|---|---|
| No nuclear signal in marrow hematopoietic cells (HPA: High) | Detection or retrieval may be insufficient (general IHC practice). | Check positive control, retrieval and antibody dilution (general IHC practice). |
| Predominantly cytoplasmic stain (UniProt Q01094: nucleus) | Nonspecific signal is possible (general IHC practice). | Compare with nuclear counterstain and antibody controls (general IHC practice). |
| Stain in skeletal muscle myocytes (HPA: Not detected) | Nonspecific binding or endogenous enzyme activity (general IHC practice). | Check reagent-omission control and enzyme blocking (general IHC practice). |
| Diffuse section background (general IHC practice) | Blocking or washing may be insufficient (general IHC practice). | Optimize blocking, washes and antibody concentration (general IHC practice). |
| Uneven staining across a section (general IHC practice) | Uneven reagent coverage or section drying (general IHC practice). | Keep sections moist and ensure even reagent coverage (general IHC practice). |
Experiment evidence
IHC & IF images showing E2F1 staining
M00257-1 · IHCMouse pancreas was stained with anti-E2F-1 rabbit antibody
M00257-1 · IHCRat pancreas was stained with anti-E2F-1 rabbit antibody
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000101412-E2F1/tissue/cerebral+cortex#imid_4979872
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000101412-E2F1/tissue/colon#imid_4979782
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000101412-E2F1/tissue/liver#imid_4979783
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000101412-E2F1/tissue/kidney#imid_4979784
HPA · ICC-IF CACO-2Human Protein Atlas immunofluorescence, CACO-2, antibody HPA029735; green = E2F1, blue = nucleus, red = microtubules.
HPA · ICC-IF CACO-2Human Protein Atlas immunofluorescence, CACO-2, antibody HPA029735; green = E2F1, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for E2F1 IHC & IF
🧪Run bone marrow first; hematopoietic cells should show E2F1 staining (HPA: High in bone marrow hematopoietic cells). Use skeletal muscle myocytes as the negative tissue (HPA: Not detected in skeletal muscle myocytes); within the bone marrow slide, cells without nuclear signal should show counterstain alone, but their absence of E2F1 should not be assumed from the HPA tissue rating (UniProt Q01094: nucleus; HPA: High in bone marrow hematopoietic cells).
Positive control tissue: Adrenal gland (Glandular cells, HPA High)
Negative control tissue: Kidney (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show E2F1 in CACO-2, Rh30, U2OS, A-431, U-251MG, with annotated localisation: Nucleoplasm (supported) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control and an isotype control matched to the primary antibody’s host species and clonality (standard IHC practice). Use E2F1 knockout material as a biological negative, and quench endogenous peroxidase in bone marrow before chromogenic detection (standard IHC practice).
⚠️Feasibility: A target-specific fixation window and the effect of fixation on E2F1 staining are unreported in the supplied evidence; the selected M00257-1 mouse-pancreas IHC caption does not state a fixative (selected-SKU caption). Antigen-retrieval dependence is unreported, so optimize retrieval for the chosen paraffin-section assay (standard IHC practice). The supplied evidence does not establish whether frozen sections or IF are easier; bone marrow requires attention to endogenous peroxidase during chromogenic detection (standard IHC practice).
HPA tissue IHC evidence for E2F1
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Enhanced — Medium consistency between antibody staining and RNA expression data.). Rows are taken directly from the HPA tissue chart — click any row's HPA link to view the source.
Positive expression · recommended positive controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adrenal gland | Glandular cells | High | Protein (IHC) | HPA → |
| Appendix | Lymphoid tissue | High | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | High | Protein (IHC) | HPA → |
| Cerebellum | Cells in molecular layer | High | Protein (IHC) | HPA → |
| Cerebral cortex | Glial cells | High | Protein (IHC) | HPA → |
Section 3
Advanced E2F1 IHC Tips
Troubleshoot E2F1 staining by assessing nuclear contrast, tissue morphology and cell type alongside antibody controls (UniProt Q01094; HPA tissue IHC; standard IHC practice).
How should I optimize retrieval when E2F1 nuclear staining is weak?
Use Tris-EDTA pH 9.0 heat-induced epitope retrieval at 95–98 °C for 20 min before staining paraffin sections (page retrieval setting). Judge retrieval by crisp nuclear contrast in intact cells, consistent with E2F1’s annotated nuclear location (UniProt Q01094; standard IHC practice). Compare a retrieved section with an unretrieved control while keeping antibody incubation and chromogen development identical, and inspect both signal and morphology (standard IHC practice). If signal remains weak, adjust heating time around the page setting or test another buffer as a documented fallback, recording whether background rises or tissue detail deteriorates (standard IHC practice).
Could fixation explain weak or patchy E2F1 staining in paraffin sections?
Target-specific fixation sensitivity is unknown from the supplied evidence, so a weak E2F1 result cannot be assigned to a particular fixative or fixation duration (evidence scope). Record the fixative, duration, processing history and section age for each specimen, then compare sections processed together under the same retrieval and staining conditions (standard IHC practice). Examine whether weak nuclear signal follows a processing batch or affects only damaged tissue; assess morphology before changing antibody concentration (standard IHC practice; UniProt Q01094 nucleus). Use a consistently processed control section in each run, and change one preparation variable at a time so any improvement can be interpreted (standard IHC practice).
What should I do if E2F1 staining appears mainly cytoplasmic?
Expect predominantly nuclear staining because E2F1 is annotated in the nucleus and HPA reports nuclear expression across most tissues (UniProt Q01094; HPA tissue IHC). HPA places E2F1 mainly in the nucleoplasm, while its additional centrosome assignment is uncertain; isolated extranuclear puncta therefore need independent confirmation (HPA subcellular). Check whether nuclear counterstain and chromogen align within intact cells, then compare the pattern with a primary-omission control and a suitably stained control section (standard IHC practice). If diffuse cytoplasmic color persists, review retrieval, antibody concentration, washing and chromogen development before counting those cells as E2F1 positive (standard IHC practice).
How can epitope accessibility affect E2F1 IHC results?
The supplied record lists one E2F1 chain spanning residues 1–437 and annotates no isoforms, so an isoform-specific explanation lacks support here (UniProt Q01094). It does list acetylation, methylation and phosphorylation sites, but their presence alone does not establish that any particular antibody loses binding after processing (UniProt Q01094; evidence scope). Check the catalog antibody’s stated immunogen or epitope, if available, before interpreting discordance between antibodies as a biological difference (standard IHC practice). For an uncertain epitope, compare independently validated antibodies and controls on matched sections, scoring nuclear localization and tissue morphology under identical development conditions (standard IHC practice; UniProt Q01094 nucleus).
How should I investigate E2F1 staining with multiplex IF or ICC?
For IF or ICC, assess E2F1 against a nuclear counterstain because its main reported location is the nucleoplasm (HPA subcellular; standard IF practice). Multiplex with a validated marker for the cell type under study, and inspect single-label controls to separate overlapping fluorescence from apparent E2F1 colocalization (standard IF practice). Choose a fluorophore channel with low tissue autofluorescence and check an unstained sample before assigning faint signal, especially when comparing fields (standard IF practice). E2F1 has no annotated transmembrane segment, so permeabilize fixed cells to expose its intracellular nuclear epitope; optimize that step without assuming an E2F1-specific fixation response (UniProt Q01094 topology; standard IF practice; evidence scope).
How can I reduce widespread brown background without losing nuclear E2F1 signal?
First compare stained sections with primary-omission and detection-only controls to locate background arising from the detection workflow (standard IHC practice). In peroxidase-based chromogenic IHC, check the peroxidase block, blocking step, washes and DAB development before changing retrieval or antibody concentration (standard IHC practice). Review intact nuclei separately from folds, damaged edges and pigment, since E2F1 is annotated as nuclear and these features can distort apparent localization (UniProt Q01094; standard IHC practice). If background persists, titrate the primary antibody and shorten chromogen development in matched sections, retaining a control that still shows interpretable nuclear contrast (standard IHC practice).
What should I measure when comparing E2F1 IHC across sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and region of interest before scoring, then count E2F1-positive nuclei rather than all brown pixels (UniProt Q01094 nucleus; standard IHC practice). Report either the percentage of positive nuclei or an H-score using intensity grades 0–3, with the H-score ranging from 0–300 (standard IHC practice). Normalize positive counts to the total nuclei in the same population, or report positive-cell density per mm² of evaluable tissue when area is the denominator (standard IHC practice). Apply identical positivity thresholds and exclude folds, necrosis and section edges consistently; record staining batch and control performance alongside results (standard IHC practice).
How can I distinguish true E2F1 signal from staining artefact?
Prioritize staining within intact nuclei, matching E2F1’s nuclear annotation and HPA’s predominantly nuclear tissue pattern (UniProt Q01094; HPA tissue IHC). Compare cell types within the section: HPA reports high staining in bone-marrow hematopoietic cells but no detection in pancreatic exocrine glandular cells, providing context rather than a universal control (HPA tissue IHC). Treat cytoplasmic haze, staining confined to cut edges or necrotic areas, and color reproduced in primary-omission controls as suspect (standard IHC practice). Check endogenous peroxidase contribution with the appropriate detection control, and interpret the selected mouse-pancreas image cautiously because its caption does not state a fixative (standard IHC practice; M00257-1 caption).
Boster reagents
Best E2F1 / Transcription factor E2F1 IHC Antibodies
E2F1 IHC images show mouse and rat pancreas (M00257-1 image captions); IF imaging shows HeLa cells (M00257 image caption). Both antibodies list human, mouse, and rat reactivity (catalog reactivity).
Real IHC data
Anti-E2F-1 Rabbit Monoclonal Antibody
Cat # M00257-1
The rendered SKU, M00257-1, has IHC images of mouse and rat pancreas (M00257-1 image captions). It lists IHC and IF applications and human, mouse, and rat reactivity, but has no IF image in the payload (M00257-1 catalog applications, reactivity, and image alts).
Which to pick: Choose M00257-1 for tissue IHC when the mouse or rat pancreas images are relevant; those captions do not report section processing or fixative (M00257-1 image captions). For IF/ICC, consider M00257: it lists both applications and has an IF image caption identifying HeLa cells; its IHC caption identifies paraffin-embedded human ovary carcinoma, with fixative unreported (M00257 catalog applications and image captions). Both are rabbit monoclonal antibodies listing human, mouse, and rat reactivity for cross-species planning (catalog titles and reactivity).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q01094 (E2F1_HUMAN, Transcription factor E2F1).
- Human Protein Atlas. E2F1 tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. E2F1 subcellular location (ICC-IF): Mainly localized to the nucleoplasm. In addition localized to the centrosome..
- Human Protein Atlas. E2F1 antibody validation summary (4 antibodies).
- U2 small nuclear RNA auxiliary factor 2, transcriptionally activated by the transcription factor Dp-1/E2F transcription factor 1 complex, enhances the growth and aerobic glycolysis of leiomyosarcoma cells. Bioengineered 2022 — PMC9278431.
- Adenovirus E2F1 overexpression sensitizes LNCaP and PC3 prostate tumor cells to radiation in vivo. International journal of radiation oncology, biology, physics 2011 — PMC4299462.
- E2F-1 targets miR-519d to regulate the expression of the ras homolog gene family member C. Oncotarget 2017 — PMC5362443.
- CircMYBL2 facilitates hepatocellular carcinoma progression by regulating E2F1 expression. Oncology research 2024 — PMC11136682.
- PubMed PMID:1638634 — UniProt-cited evidence.
- PubMed PMID:1638635 — UniProt-cited evidence.
- PubMed PMID:1448092 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for E2F1 (Transcription factor E2F1). Expected localisation Nuclear staining in most tissues (HPA tissue IHC). Antigen retrieval: Tris-EDTA pH 9.0 HIER, 95–98 °C, 20 min (rule: nuclear antigen). Validated IHC antibody: M00257-1 at 1:200-1:1000 (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Glandular cells); negative Kidney. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Tissue sections; selected-image fixative not specified (standard IHC workflow); 2. Fixation: Image fixative and duration unreported (datasheet M00257-1); 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-E2F1, 1:200-1:1000 (datasheet M00257-1); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across sections (standard IHC practice; not target-specific); Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image M00257-1); Caveat: Glomerular cells have no detected signal (HPA tissue IHC); Isoform / epitope: 0 annotated isoforms; no processing (UniProt); Regulation: G1–S activity; abundance regulation unreported (UniProt). Evidence: UniProt Q01094, HPA tissue IHC · Enhanced, HPA ICC-IF · Mainly localized to the nucleoplasm. In addition localized to the…, Datasheet M00257-1, PMC5362443, PMC11136682. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/E2F1/immunohistochemistry-ihc-guide.