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ZBTB4 / Zinc finger and BTB domain-containing protein 4 · IHC design guide
Design Immunohistochemistry for ZBTB4
Plan ZBTB4 chromogenic IHC around its predominantly nuclear tissue staining (HPA tissue IHC). Start antibody A08155 at 2.5 μg/mL (datasheet), and compare decidual cells with appendix glandular cells, where staining was high and undetected, respectively (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
ZBTB4 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 | Predominantly nuclear; lesser cytoplasmic staining (HPA tissue IHC) | |
| Staining pattern | Most tissues show nuclear and lesser cytoplasmic staining (HPA tissue IHC) | |
| Antigen retrieval | Tris-EDTA pH 9.0 HIER, 95–98 °C, 20 min (rule: nuclear antigen) | |
| Positive control | Placenta+4 more · see all | |
| Negative control | Appendix+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep paraffin-section fixation consistent across samples. (standard IHC practice; not target-specific) | |
| Caveat | Staining–RNA consistency is medium; verify specificity (HPA tissue IHC) | |
| Regulation | Low tissue specificity; no regulator annotated (HPA RNA; UniProt) | |
| Isoform / epitope | No isoforms annotated; one chain spans residues 1–1013 (UniProt) |
Section 1
Recommended ZBTB4 IHC & IF Protocols
Compare the catalog antibody’s IHC-P protocol with four published ZBTB4 IHC protocols (PMC10035130; PMC7547138; PMC9941281; PMC10875348).
| Sample | Tissue sections; selected-image fixative not specified (standard IHC workflow) |
| Fixation | Image fixative and duration unreported (datasheet A08155); 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-ZBTB4, 2.5 μg/mL (datasheet A08155) |
| 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 | ZBTB4-positive staining in decidual cells of placenta (HPA tissue IHC: High). HPA tissue profile: Nuclear and to a lesser extent cytoplasmic expression in most tissues. No signal in the no-primary control. |
💡Decision noteStart with Tris-EDTA pH 9.0 HIER at 95–98 °C for 20 min (page retrieval rule: nuclear antigen). Citrate heat retrieval is reported for colorectal tissue (PMC7547138).
Section 2
What Is the Expected ZBTB4 Staining Pattern?
ZBTB4 should stain mainly nuclei in paraffin sections, with weaker cytoplasmic staining possible across many tissues (HPA tissue IHC: Supported). Strong staining is reported in placental decidual cells; several other listed cell types show medium staining (HPA tissue IHC). Its nuclear and chromocenter localization, with no transmembrane segment, supports a nuclear interpretation (UniProt Q9P1Z0). HPA reports medium agreement with RNA data and says external verification is pending (HPA tissue IHC).
| Decidual cell nuclei stain strongly; adipocyte, adrenal glandular, or breast glandular cell nuclei stain moderately. | This fits the reported high placental decidual signal and medium signal in the listed cell types (HPA tissue IHC). Judge staining in the named cells and their nuclei, since the tissue label alone does not identify which cells should be positive (HPA tissue IHC). |
| Signal is predominantly membranous, or cytoplasm stains much more strongly than nuclei across the section. | That distribution conflicts with the reported mainly nuclear IHC pattern and the lack of a transmembrane segment (HPA tissue IHC; UniProt Q9P1Z0 topology). Limited cytoplasmic signal can occur, so assess whether a convincing nuclear pattern remains before calling the stain discordant (HPA tissue IHC). |
| Appendix or duodenal glandular cells, or liver cholangiocytes, stain strongly. | Those specific cell populations were reported as not detected (HPA tissue IHC). Check cell identity and controls, then consider nonspecific antibody binding or endogenous chromogen activity as possible explanations (standard IHC practice). A negative designation for these cells does not classify every cell in the section as negative (HPA tissue IHC). |
| Color is spread across tissue or present in blank regions without recognizable nuclear boundaries. | This is difficult to score as ZBTB4 because the expected pattern is mainly nuclear (HPA tissue IHC). Uneven blocking, excessive antibody concentration, or detection background are general IHC possibilities; they are not established ZBTB4-specific effects (standard IHC practice). |
| Placental decidual cells show no detectable nuclear staining. | The result disagrees with the reported high staining in those cells (HPA tissue IHC). Confirm that decidual cells are represented and examine the IHC run controls and catalog antibody conditions before interpreting the absence as biological (standard IHC practice). HPA's Supported rating still awaits external verification (HPA tissue IHC). |
💡Expected ZBTB4 appearanceCall a section consistent with ZBTB4 when identifiable placental decidual cells show strong, predominantly nuclear chromogenic signal, allowing lesser cytoplasmic signal; strong membranous or uniformly diffuse staining is suspect (HPA tissue IHC; UniProt Q9P1Z0 topology).
| Cell population chosen for assessment | Placental decidual cells offer a reported high-signal reference; adipocytes and several listed glandular or neural cells have medium signal (HPA tissue IHC). Appendix and duodenal glandular cells, liver cholangiocytes, and splenic red-pulp cells are reported not detected; evaluate the specified cells rather than labeling whole organs positive or negative (HPA tissue IHC). |
| Expected compartment | UniProt places ZBTB4 in the nucleus and at chromocenters; tissue IHC reports mainly nuclear with lesser cytoplasmic staining (UniProt Q9P1Z0; HPA tissue IHC). Use nuclear predominance to interpret chromogenic sections. Nuclear bodies are an ICC-IF observation and should not be required as a resolvable chromogenic IHC feature (HPA subcellular ICC-IF). |
| Strength of the tissue evidence | The tissue profile is rated Supported, with medium agreement between staining and RNA expression and external verification pending (HPA tissue IHC). Two listed antibodies have Supported IHC status, but the payload does not assign Enhanced IHC validation (HPA antibodies). Treat departures from the pattern as findings to check, especially when based on a single section (standard IHC practice). |
| Topology and processing | ZBTB4 has no transmembrane segment or signal peptide, and the annotated chain spans residues 1–1013 (UniProt Q9P1Z0). These annotations support interpreting a mainly nuclear result rather than expecting a membrane rim; they do not predict IHC sensitivity or explain a failed stain (UniProt Q9P1Z0; HPA tissue IHC). |
| Fixation and epitope uncertainty | The supplied sources give no ZBTB4-specific fixation response or antibody epitope position (UniProt Q9P1Z0; HPA tissue IHC). Follow the catalog antibody's validated IHC-P conditions when available and record retrieval conditions for comparison; do not infer a ZBTB4-specific retrieval requirement from tissue staining levels (standard IHC practice). |
| Situation | Likely cause | Next action |
|---|---|---|
| No nuclear signal in placental decidual cells. | The observed result conflicts with their reported high staining; missed cell identification or an IHC run issue is possible (HPA tissue IHC; standard IHC practice). | Confirm decidual cells on morphology, inspect run controls, and check that the IHC-validated antibody and its stated IHC-P conditions were used (standard IHC practice). |
| Signal is faint in adipocytes or listed medium-staining glandular cells. | Their reported level is medium, so a weaker result than placental decidual cells can fit the profile (HPA tissue IHC). | Score nuclear signal in the named cells against a concurrent positive reference before changing conditions; document section-to-section variation (HPA tissue IHC; standard IHC practice). |
| Strong signal appears in appendix or duodenal glandular cells. | Those cells were reported not detected; nonspecific staining or endogenous detection activity is possible (HPA tissue IHC; standard IHC practice). | Verify the cell population, inspect no-primary and detection controls, and review blocking and detection steps if background persists (standard IHC practice). |
| Most staining outlines membranes or fills cytoplasm, with little nuclear signal. | This disagrees with mainly nuclear tissue IHC and nuclear UniProt localization; lesser cytoplasmic staining alone remains compatible (HPA tissue IHC; UniProt Q9P1Z0). | Compare nuclear and cytoplasmic signal in the same named cells, then inspect antibody concentration and detection controls for nonspecific signal (standard IHC practice). |
| Diffuse chromogen obscures nuclei across positive and negative regions. | General IHC background can prevent compartment scoring; the supplied sources do not establish a ZBTB4-specific fixation or retrieval cause (standard IHC practice; HPA tissue IHC). | Review no-primary controls, blocking, antibody dilution, washes, and detection timing under the validated IHC-P workflow; reassess identifiable nuclei afterward (standard IHC practice). |
| An IF image shows nuclear bodies or some cytosolic signal that the IHC slide does not resolve. | Nuclear bodies and nucleoplasm are supported ICC-IF locations; cytosol is uncertain, while tissue IHC describes mainly nuclear staining (HPA subcellular ICC-IF; HPA tissue IHC). | Interpret the chromogenic IHC section by its reported tissue and cell pattern. Use the separate IF/ICC guide for fluorescence-specific assessment (HPA tissue IHC; HPA subcellular ICC-IF). |
Experiment evidence
IHC & IF images showing ZBTB4 staining
A08155 · IHCImmunohistochemistry of ZBTB4 in human brain tissue with ZBTB4 antibody at 2.5 μg/mL.
A08155 · IFImmunofluorescence of ZBTB4 in human brain tissue with ZBTB4 antibody at 20 μg/mL.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000174282-ZBTB4/tissue/cerebral+cortex#imid_17822987
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000174282-ZBTB4/tissue/colon#imid_17822929
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000174282-ZBTB4/tissue/liver#imid_17822930
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000174282-ZBTB4/tissue/kidney#imid_17822923
HPA · ICC-IF HaCaTHuman Protein Atlas immunofluorescence, HaCaT, antibody HPA064763; green = ZBTB4, blue = nucleus, red = microtubules.
HPA · ICC-IF HaCaTHuman Protein Atlas immunofluorescence, HaCaT, antibody HPA064763; green = ZBTB4, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for ZBTB4 IHC & IF
🧪Run placenta first: decidual cells should stain (HPA: High in decidual cells). Use appendix glandular cells as the negative tissue (HPA: Not detected); on the placenta slide, compare with neighboring cells that lack nuclear staining, without assuming a specific neighboring cell type is negative.
Positive control tissue: Placenta (Decidual cells, HPA High)
Negative control tissue: Appendix (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show ZBTB4 in HaCaT, U-251MG, U2OS, with annotated localisation: Nuclear bodies (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 immunoglobulin class; use ZBTB4 knockout material as a biological negative (standard IHC practice). Block endogenous peroxidase and inspect blood-rich areas of the placenta for chromogenic background (standard chromogenic IHC practice).
⚠️Feasibility: A target-specific fixation window and antigen retrieval requirement are unreported in the supplied evidence; optimize retrieval empirically. The selected A08155 tissue-IHC caption does not state a fixative (caption: fixative unreported). The evidence does not establish whether frozen sections or IF are easier than paraffin IHC; placental blood may contribute peroxidase background (standard chromogenic IHC practice).
HPA tissue IHC evidence for ZBTB4
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Supported — 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 |
|---|---|---|---|---|
| Placenta | Decidual cells | High | Protein (IHC) | HPA → |
| Adipose tissue | Adipocytes | Medium | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | Medium | Protein (IHC) | HPA → |
| Breast | Glandular cells | Medium | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | Medium | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Appendix | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Duodenum | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Liver | Cholangiocytes | Not detected | Protein (IHC) | HPA → |
| Small intestine | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Spleen | Cells in red pulp | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced ZBTB4 IHC Tips
Troubleshoot ZBTB4 staining in chromogenic paraffin section IHC, using nuclear localisation and tissue patterns to guide controls and interpretation.
How should I retrieve ZBTB4 when nuclear staining is weak?
Start with Tris-EDTA pH 9.0 heat-induced antigen retrieval at 95–98 °C for 20 min (page retrieval rule: nuclear antigen). Let sections cool in retrieval buffer, then compare staining with a consistently processed positive control; the expected signal is predominantly nuclear (UniProt: nucleus and chromocenters; HPA tissue IHC: mainly nuclear). If staining remains weak, test a shorter or longer heating period on adjacent sections while keeping antibody concentration and detection constant (standard IHC practice). Judge improvement by nuclear signal and preserved tissue morphology, since stronger diffuse cytoplasmic colour would not establish better ZBTB4 detection (HPA tissue IHC: lesser cytoplasmic expression).
Could fixation explain inconsistent ZBTB4 staining between paraffin blocks?
Target-specific ZBTB4 sensitivity to fixation is unknown from the supplied evidence; the catalog antibody’s human brain IHC caption does not state a fixative (A08155 tissue-IHC caption). Record each block’s fixative and fixation duration, and compare sections processed with the same retrieval and detection conditions before attributing differences to the antibody (standard IHC practice). Review morphology for overfixation, underfixation or processing damage, and include a similarly processed positive tissue section in each run (standard IHC practice). Do not infer fixation tolerance from the reported tissue staining pattern or ZBTB4’s nuclear localisation and phosphorylation sites (HPA tissue IHC; UniProt: localisation and modified residues).
Where should convincing ZBTB4 staining appear in a paraffin section?
Prioritise nuclear staining, which matches ZBTB4’s reported nucleus and chromocenter localisation (UniProt: subcellular location). Nuclear bodies are the supported main location in cell imaging, with supported nucleoplasmic and uncertain cytosolic localisation (HPA subcellular); chromogenic sections may not resolve individual nuclear bodies (standard IHC practice). HPA tissue IHC reports predominantly nuclear staining with lesser cytoplasmic expression across most tissues, so score nuclear and cytoplasmic signals separately (HPA tissue IHC). Compare the staining pattern with intact nuclei and neighbouring cells on the same section, and investigate diffuse cytoplasmic colour before calling it ZBTB4 positive (standard IHC practice; HPA tissue IHC: mainly nuclear).
Could epitope choice explain discrepant ZBTB4 IHC results?
Check the antibody’s stated immunogen or epitope against the full 1–1013 amino acid ZBTB4 chain; the supplied record annotates 0 isoforms but does not locate this antibody’s epitope (UniProt: chain and isoforms; supplied antibody evidence). ZBTB4 contains a BTB domain at residues 30–152 and reported phosphorylation at residues including 391, 795, 797 and 983 (UniProt: domains and modified residues). Epitope masking remains a possibility to test, not an established explanation for this antibody (standard IHC practice; supplied antibody evidence). Compare adjacent sections under controlled retrieval conditions and seek agreement in nuclear localisation before interpreting different staining patterns biologically (UniProt: nucleus; standard IHC practice).
How can I check ZBTB4 localisation by multiplex IF?
For a separate IF experiment, pair ZBTB4 with a validated marker for the expected cell population and a nuclear counterstain, then assess nuclear overlap within individual cells (HPA tissue IHC: cell-specific patterns; UniProt: nucleus). Select a fluorophore in a channel with low measured tissue autofluorescence, using unstained and single-stain controls to check bleed-through (standard IF practice). Because ZBTB4 has no transmembrane segment and is predominantly nuclear, permeabilise cells or sections sufficiently for intracellular antibody access while preserving nuclear structure (UniProt: topology and localisation; standard IF practice). HPA supports nuclear bodies and nucleoplasm in cell imaging, whereas its cytosolic assignment is uncertain, so treat isolated cytosolic fluorescence cautiously (HPA subcellular).
What should I check when ZBTB4 IHC has diffuse background?
First inspect a no-primary control and the section edges to distinguish detection background from antibody-associated staining (standard IHC practice). Block endogenous peroxidase before chromogenic development and titrate primary antibody, detection reagent and DAB exposure on comparable sections (standard chromogenic IHC practice). The catalog antibody’s human brain image reports 2.5 µg/mL, but its caption does not establish an optimal concentration for every tissue or disclose fixation (A08155 tissue-IHC caption). Retain conditions that reveal interpretable nuclear signal without broad haze, and assess cytoplasmic colour separately because tissue IHC reports lesser cytoplasmic ZBTB4 expression (HPA tissue IHC: localisation).
How should I quantify ZBTB4 across stained tissue sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and tissue region before scoring, then record the percentage of positive nuclei and an intensity-based nuclear H-score of 0–300 (standard IHC scoring practice; UniProt: nuclear localisation). Use identical thresholds, section thickness, imaging settings and chromogenic development conditions across the comparison set (standard IHC practice). Normalise positive nuclei to the number of evaluable nuclei in each region; if spatial abundance matters, also report positive nuclei per mm² of viable tissue (standard IHC quantification practice). Keep cytoplasmic staining as a separate measure, since HPA describes predominantly nuclear and lesser cytoplasmic tissue expression (HPA tissue IHC).
When is an apparent ZBTB4-positive cell likely to be an artefact?
A convincing call should show signal in an intact nucleus and a plausible cell population, consistent with ZBTB4’s nuclear localisation and the cell-specific tissue patterns reported by HPA (UniProt: localisation; HPA tissue IHC). Scrutinise staining restricted to cut edges, folds or necrotic regions, and compare it with intact tissue away from those areas (standard IHC practice). Diffuse colour in the wrong compartment, or colour persisting in a no-primary control, warrants checking nonspecific detection and endogenous enzyme activity (standard chromogenic IHC practice). HPA rates its tissue IHC evidence Supported but pending external verification, so corroborate unexpected patterns with an independent control (HPA tissue IHC: reliability).
Boster reagents
Best ZBTB4 / Zinc finger and BTB domain-containing protein 4 IHC Antibodies
A08155 has human brain IHC and IF images (catalog image captions), with Human and Mouse reactivity listed (catalog: reactivity).
Real IHC data
Anti-ZBTB4 Antibody
Cat # A08155
A08155 is listed for IHC-P and IF (catalog: applications). Its images show ZBTB4 staining in human brain tissue at 2.5 μg/mL for IHC and 20 μg/mL for IF (catalog: image captions).
Which to pick: For paraffin-section IHC, choose A08155 because IHC-P is listed (catalog: applications) and its own IHC image shows human brain tissue at 2.5 μg/mL (catalog: IHC image caption); the fixative is unreported (catalog: IHC image caption). For IF, choose A08155 based on its human brain IF image at 20 μg/mL (catalog: IF image caption); ICC validation is unreported (catalog: applications and image captions). For mouse samples, A08155 lists Mouse reactivity (catalog: reactivity), while the supplied IHC and IF images show human tissue (catalog: image captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q9P1Z0 (ZBTB4_HUMAN, Zinc finger and BTB domain-containing protein 4).
- Human Protein Atlas. ZBTB4 tissue IHC expression (reliability: Supported).
- Human Protein Atlas. ZBTB4 subcellular location (ICC-IF): Mainly localized to the nuclear bodies. In addition localized to the nucleoplasm and cytosol..
- Human Protein Atlas. ZBTB4 antibody validation summary (2 antibodies).
- Identification of ZBTB4 as an immunological biomarker that can inhibit the proliferation and invasion of pancreatic cancer. BMC cancer 2023 — PMC10035130.
- Expression of Zinc Finger and BTB Domain-Containing 4 in Colorectal Cancer and Its Clinical Significance. Cancer management and research 2020 — PMC7547138.
- miRNA-576-5p promotes endometrial cancer cell growth and metastasis by targeting ZBTB4. Clinical & translational oncology : official publication of the Federation of Spanish Oncology Societies and of the National Cancer Institute of Mexico 2023 — PMC9941281.
- Demethylase FTO inhibits the occurrence and development of triple-negative breast cancer by blocking m (6)A-dependent miR-17-5p maturation-induced ZBTB4 depletion. Acta biochimica et biophysica Sinica 2024 — PMC10875348.
- PubMed PMID:10819331 — UniProt-cited evidence.
- PubMed PMID:12168954 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for ZBTB4 (Zinc finger and BTB domain-containing protein 4). Expected localisation Predominantly nuclear; lesser cytoplasmic staining (HPA tissue IHC). Antigen retrieval: Tris-EDTA pH 9.0 HIER, 95–98 °C, 20 min (rule: nuclear antigen). Validated IHC antibody: A08155 at 2.5 μg/mL (the product's own IHC validation image is shown). Controls (HPA): positive Placenta (Decidual cells); negative Appendix. 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 A08155); 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-ZBTB4, 2.5 μg/mL (datasheet A08155); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep paraffin-section fixation consistent across samples. (standard IHC practice; not target-specific); Caveat: Staining–RNA consistency is medium; verify specificity (HPA tissue IHC); Isoform / epitope: No isoforms annotated; one chain spans residues 1–1013 (UniProt); Regulation: Low tissue specificity; no regulator annotated (HPA RNA; UniProt). Evidence: UniProt Q9P1Z0, HPA tissue IHC · Supported, HPA ICC-IF · Mainly localized to the nuclear bodies. In addition localized to the…, Datasheet A08155, PMC10035130, PMC7547138. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/ZBTB4/immunohistochemistry-ihc-guide.