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HOPX / Homeodomain-only protein · IHC design guide
Design Immunohistochemistry for HOPX
Plan HOPX IHC in paraffin sections using the human-reactive A05019 antibody, starting at 5 μg/mL (datasheet: A05019). Compare nuclear and cytoplasmic staining with tissue patterns, including high staining in skin granular-layer cells (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
HOPX 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 and cytoplasmic staining (HPA tissue IHC) | |
| Staining pattern | Nuclear/cytoplasmic in squamous cells, glands and pneumocytes (HPA tissue IHC) | |
| Antigen retrieval | Tris-EDTA pH 9.0 HIER, 95–98 °C, 20 min (rule: nuclear antigen) | |
| Positive control | Duodenum+4 more · see all | |
| Negative control | Adipose tissue+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A05019) | |
| Caveat | Staining varies by cell population within a tissue (HPA tissue IHC) | |
| Regulation | Down-regulated in some cancers (UniProt) | |
| Isoform / epitope | 4 isoforms; epitope coverage is unspecified (UniProt) |
Section 1
Recommended HOPX IHC & IF Protocols
Compare the catalog antibody's IHC-P protocol with published chromogenic HOPX IHC protocols for thyroid, pancreatic, and head and neck tissues (PMC6800262; PMC3488580; PMC5146930).
| Sample | Tissue sections; selected-image fixative not specified (standard IHC workflow) |
| Fixation | Image fixative and duration unreported (datasheet A05019); 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-HOPX, 5 μg/mL (datasheet A05019) |
| 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 | HOPX-positive staining in endocrine cells of duodenum (HPA tissue IHC: High). HPA tissue profile: Nuclear and cytoplasmic expression in squamous epithelia, glandular cells of thyroid and epididymis and in pneumocytes. No signal in the no-primary control. |
💡Decision noteStart with Tris-EDTA pH 9.0 at 95–98 °C for 20 min (page retrieval rule); citrate pH 6.0 is a published alternative (PMC6800262).
Section 2
What Is the Expected HOPX Staining Pattern?
HOPX should appear in nuclei and cytoplasm, with staining interpreted by cell type rather than as a uniform tissue-wide signal (UniProt Q9BPY8; HPA tissue IHC). HPA reports nuclear and cytoplasmic staining in squamous epithelia, thyroid and epididymal glandular cells, and pneumocytes; its tissue IHC reliability is Enhanced, with medium consistency between staining and RNA expression (HPA tissue IHC). HOPX has no transmembrane segment (UniProt Q9BPY8 topology).
| Distinct nuclear staining, with possible cytoplasmic staining, appears in skin granular-layer cells. | This fits a high-staining cell population (HPA: High in skin granular-layer cells) and the documented nuclear and cytoplasmic locations (HPA tissue IHC; UniProt Q9BPY8). Judge the relevant cells rather than the darkness of the whole section. |
| A sharply outlined, membrane-only signal dominates otherwise unstained skin epithelial cells. | That is atypical for the expected nuclear and cytoplasmic pattern (HPA tissue IHC; UniProt Q9BPY8). Check staining specificity and detection artifacts before scoring it as HOPX. Interpret other tissues separately: HPA labels cerebellar granular-cell staining as cytoplasm/membrane (HPA: Medium in cerebellum). |
| Strong staining appears mainly in bronchial respiratory epithelial cells or adipocytes. | These cell populations are listed as not detected (HPA: bronchus respiratory epithelial cells; adipose tissue adipocytes). Consider cross-reactivity or endogenous detection activity, and compare the same run with a known-positive section. A single unexpected cell population does not establish a new HOPX pattern. |
| Color spreads across stroma, empty spaces, and many unrelated cells without clear cellular boundaries. | A diffuse field cannot be assigned confidently to the documented nuclear and cytoplasmic locations (HPA tissue IHC; UniProt Q9BPY8). In general chromogenic IHC practice, incomplete blocking, residual detection reagent, and inadequate washing can create background; inspect controls before interpreting weak cells. |
| A skin section lacks staining in its granular-layer cells. | HPA reports high staining in those cells (HPA: High in skin granular layer). First assess section quality, retrieval and detection controls under the assay's validated conditions. Absence in one run may reflect technical failure; it cannot alone establish absent HOPX expression. |
💡Expected HOPX appearanceCall a positive result when the relevant cells show discernible nuclear staining, with possible cytoplasmic staining—for example, high staining in skin granular-layer cells (HPA tissue IHC; HPA: High in skin); diffuse field color or a dominant membrane-only outline is suspect in that setting (UniProt Q9BPY8 topology; general IHC interpretation).
| Cell population and tissue | Expected intensity changes by cell population: duodenal and small-intestinal endocrine cells are High, epididymal glandular cells are Medium, and bronchial respiratory epithelial cells are Not detected (HPA tissue IHC). Select and score controls at the cell level. |
| Compartment and topology | UniProt lists nucleus and cytoplasm and no transmembrane segment (UniProt Q9BPY8). HPA tissue IHC reports both compartments; compartment alone is insufficient without the expected cell type (HPA tissue IHC). |
| Antibody evidence | HPA030180 and CAB018632 each have Enhanced IHC status (HPA antibody validation). HPA describes tissue IHC reliability as Enhanced but reports only medium consistency with RNA expression (HPA tissue IHC); retain that qualification when judging an unexpected pattern. |
| Isoforms and processing | Four isoforms are listed, while the supplied record gives no isoform-specific IHC pattern (UniProt Q9BPY8). It lists a 1–73 chain, no signal peptide or propeptide, and no annotated glycosylation sites (UniProt Q9BPY8); these facts do not predict retrieval behavior. |
| IF/ICC: where should signal appear? | Mainly in nuclear bodies, with additional cytosolic signal (HPA ICC-IF: approved locations). That observation answers the localisation question for IF/ICC; use its separate guide for assay setup. It does not override the tissue-specific chromogenic IHC pattern (HPA tissue IHC). |
| Situation | Likely cause | Next action |
|---|---|---|
| No color in granular-layer cells of a skin positive control. | The expected population is High (HPA: skin granular layer); a failed control leaves staining sensitivity unresolved. | Check the run's reagent and detection controls, then review section integrity, retrieval conditions and primary-antibody dilution against the validated IHC procedure (general IHC practice). Do not assign a HOPX-negative result from this run. |
| Signal is nuclear but appears chiefly in cells listed as not detected. | Correct-looking localisation does not establish cell-type specificity (UniProt Q9BPY8; HPA tissue IHC). Off-target binding or endogenous detection activity is possible (general IHC practice). | Compare a known-positive tissue in the same run and inspect appropriate primary-omission and detection controls (general IHC practice). Report the unexpected cell population separately rather than treating all nuclear color as HOPX. |
| Broad brown haze obscures nuclei and cytoplasmic boundaries. | Diffuse color is difficult to reconcile with cellular HOPX localisation (HPA tissue IHC; UniProt Q9BPY8); nonspecific detection or inadequate washing can contribute (general IHC practice). | Review blocking, washes, detection-reagent exposure and negative controls under the laboratory's validated chromogenic IHC workflow (general IHC practice). Score only cells whose signal can be localised reliably. |
| A membrane-only outline is scored as positive in skin. | That appearance conflicts with the expected nuclear and cytoplasmic pattern in squamous epithelium and with the absence of a transmembrane segment (HPA tissue IHC; UniProt Q9BPY8). | Reassess morphology and background, then compare staining with the documented granular-layer pattern (HPA: High in skin granular layer). Do not apply this skin rule to cerebellar granular cells, for which HPA reports cytoplasm/membrane staining (HPA: Medium in cerebellum). |
| A low-staining or not-detected population is used as the only positive control. | Esophageal squamous epithelial cells are Low, while bronchial respiratory epithelial cells are Not detected (HPA tissue IHC). Either choice gives limited evidence that the run can reveal a High population. | Include a section containing skin granular-layer cells or duodenal endocrine cells, both reported High (HPA tissue IHC). Evaluate those specific cells before interpreting weak or absent staining in the test tissue. |
| IF/ICC nuclear-body signal is used to reject a chromogenic tissue result. | HPA's approved nuclear-body and cytosolic locations come from ICC-IF, while its tissue IHC profile describes nuclear and cytoplasmic expression (HPA ICC-IF; HPA tissue IHC). | Judge the paraffin-section result against the tissue IHC cell population and compartment evidence (HPA tissue IHC). Consult the separate IF/ICC guide when interpreting fluorescent cell images. |
Experiment evidence
IHC & IF images showing HOPX staining
A05019 · IHCImmunohistochemistry of HOPX in human heart tissue with HOPX antibody at 5 μg/mL.
A05019 · IFImmunofluorescence of HOPX in human heart tissue with HOPX antibody at 20 μg/mL.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000171476-HOPX/tissue/cerebral+cortex#imid_18413731
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000171476-HOPX/tissue/colon#imid_18413665
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000171476-HOPX/tissue/liver#imid_18413666
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000171476-HOPX/tissue/kidney#imid_18413667
HPA · ICC-IF HUVEC/TERT2Human Protein Atlas immunofluorescence, HUVEC/TERT2, antibody HPA030180; green = HOPX, blue = nucleus, red = microtubules.
HPA · ICC-IF HUVEC/TERT2Human Protein Atlas immunofluorescence, HUVEC/TERT2, antibody HPA030180; green = HOPX, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for HOPX IHC & IF
🧪Run duodenum first: its endocrine cells should stain (HPA: High in duodenum endocrine cells). Use adipose tissue as the negative, where adipocytes are not detected (HPA: Not detected in adipocytes); neighboring non-endocrine cells on the duodenum slide should be assessed for background rather than assumed to be HOPX-negative.
Positive control tissue: Duodenum (Endocrine cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show HOPX in HUVEC/TERT2, HaCaT, ASC52telo, with annotated localisation: Nuclear bodies (approved) (HPA subcellular).
Technical controls: Include no-primary (secondary-only) and host-matched control IgG, with an isotype match for a monoclonal antibody; use HOPX knockout material or peptide competition, if available, to test specificity. For chromogenic duodenum IHC, quench endogenous peroxidase and check the no-primary slide for residual tissue background.
⚠️Feasibility: The supplied evidence reports no HOPX-specific fixation window or retrieval dependence, so optimize antigen retrieval on paraffin sections. The A05019 heart IHC caption gives 5 µg/mL but does not report the fixative (A05019 caption). HPA ICC-IF images show nuclear-body and cytosolic localization (HPA: subcellular), but do not establish that IF or frozen sections are easier; in duodenum, distinguish sparse endocrine-cell staining from epithelial background (HPA: High in duodenum endocrine cells).
HPA tissue IHC evidence for HOPX
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 |
|---|---|---|---|---|
| Duodenum | Endocrine cells | High | Protein (IHC) | HPA → |
| Skin | Cells in granular layer | High | Protein (IHC) | HPA → |
| Small intestine | Endocrine cells | High | Protein (IHC) | HPA → |
| Vagina | Squamous epithelial cells | High | Protein (IHC) | HPA → |
| Cerebellum | Granular cells - cytoplasm/membrane | 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 → |
| Bone marrow | Hematopoietic 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 HOPX IHC Tips
Troubleshoot HOPX staining in paraffin sections by checking retrieval, cell identity, subcellular pattern and controls before interpreting chromogenic signal.
How should I adjust retrieval when HOPX staining is weak?
Start with Tris-EDTA at pH 9.0, heated to 95–98 °C for 20 min (page retrieval setting). If staining remains weak, compare a shorter heating time on serial sections while keeping the buffer and antibody conditions constant; review tissue preservation alongside signal intensity (standard IHC practice). Record whether any gain appears in the expected cells and nuclear or cytoplasmic compartments, since HOPX can occupy both (UniProt Q9BPY8 subcellular location; HPA tissue IHC profile). A stronger diffuse stain alone does not establish improved detection; compare it with a no-primary control and the original retrieval condition (standard IHC practice).
Could fixation explain weak or uneven HOPX staining?
The supplied HOPX evidence does not establish target-specific fixation sensitivity: the A05019 heart tissue-IHC caption gives 5 µg/mL but does not state the fixative (A05019 tissue-IHC caption). Compare sections with documented, consistent processing before changing antibody concentration, because fixation and processing can alter antigen accessibility in paraffin sections (standard IHC practice). Check for uneven staining across the section and compare morphology, retrieval exposure and staining in neighboring cells processed together (standard IHC practice). Do not infer a HOPX-specific fixation effect from its nuclear and cytoplasmic distribution or lack of annotated modifications (UniProt Q9BPY8 subcellular location and modified residues).
Should I expect nuclear or cytoplasmic HOPX staining?
Evaluate nuclear and cytoplasmic staining separately: both locations are annotated for HOPX, and tissue IHC reports both in selected cell populations (UniProt Q9BPY8 subcellular location; HPA tissue IHC profile). HPA cell imaging places the main signal in nuclear bodies, with additional cytosolic signal, but that pattern comes from ICC/IF rather than the paraffin section being scored (HPA subcellular). Use the hematoxylin counterstain to define nuclei, then assess whether chromogen is intracellular and follows intact cell boundaries (standard IHC practice). A membrane-only rim would need independent validation, since HOPX has no annotated transmembrane segment (UniProt Q9BPY8 topology).
Could isoforms or epitope masking explain discordant HOPX results?
HOPX has 4 annotated isoforms, but the supplied evidence does not identify which isoforms the IHC-validated antibody recognizes (UniProt Q9BPY8 isoforms; supplied antibody evidence). Check the antibody's stated immunogen or epitope against the isoforms being studied before treating absent staining as absent protein (standard IHC practice). Compare sections across the same retrieval condition and tissue processing batch; a change in staining after retrieval is consistent with altered epitope access, without identifying a particular isoform (standard IHC practice). HOPX has no annotated glycosylation sites or modified residues here, so do not assign a staining discrepancy to either modification without additional evidence (UniProt Q9BPY8 glycosylation and modified residues).
How can I check HOPX localisation by IF alongside IHC?
For a separate IF/ICC experiment, pair HOPX with a marker of the cell population being assessed, such as an appropriate squamous epithelial marker when investigating HPA-reported squamous staining (HPA tissue IHC profile; standard IF practice). Select fluorophores after inspecting unstained tissue, favoring channels with less autofluorescence and minimizing spectral overlap with the companion marker (standard IF practice). HOPX has no transmembrane segment and is annotated in nuclear and cytoplasmic compartments, so permeabilise cells to give antibodies access to intracellular epitopes and titrate that step against preserved morphology (UniProt Q9BPY8 topology and subcellular location; standard IF practice). Treat the resulting fluorescence pattern as a localisation check, while scoring the chromogenic paraffin sections on their own controls (standard IHC/IF practice).
What should I check when HOPX chromogen is diffuse?
Run a no-primary control and inspect whether diffuse chromogen persists; signal there points to detection chemistry or tissue background rather than primary-antibody binding (standard IHC practice). In a peroxidase and DAB workflow, check the peroxidase block, blocking step, washes and chromogen development time, adjusting 1 variable at a time (standard chromogenic IHC practice). Compare background with the expected intracellular pattern: HOPX is annotated in nucleus and cytoplasm, while HPA reports staining in particular epithelial and glandular populations (UniProt Q9BPY8 subcellular location; HPA tissue IHC profile). Review section edges and damaged areas separately before changing the antibody concentration, because local staining artefacts can mimic a stronger result (standard IHC practice).
How should I score heterogeneous HOPX staining? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, since HOPX staining can be nuclear or cytoplasmic and varies by cell type (HPA tissue IHC profile). For chromogenic sections, report the percentage of positive target cells and an H-score based on staining intensity, recording nuclear and cytoplasmic results separately when both are present (standard IHC practice). If the question concerns how many positive cells occupy an area, report positive-cell density per mm² and normalize counts to the measured viable tissue area (standard IHC practice). Keep exposure to retrieval, counterstain, threshold and region-selection rules consistent across specimens, and exclude damaged or necrotic regions by a prespecified rule (standard IHC practice).
When is apparent HOPX positivity likely to be artefactual?
A credible result places intracellular chromogen in intact cells with a plausible nuclear or cytoplasmic pattern; HOPX is annotated in both compartments (UniProt Q9BPY8 subcellular location; standard IHC practice). Check cell identity against the region being examined: HPA reports high staining in skin granular-layer cells and duodenal endocrine cells, but no detected staining in adipocytes from adipose tissue (HPA tissue IHC). Treat membrane-only outlines, section-edge enrichment and staining confined to necrotic areas cautiously, then compare them with tissue morphology and a no-primary control (UniProt Q9BPY8 topology; standard IHC practice). In peroxidase detection, persistent no-primary chromogen also warrants checking endogenous enzyme activity before attributing the signal to HOPX (standard chromogenic IHC practice).
Boster reagents
Best HOPX / Homeodomain-only protein IHC Antibodies
A05019 has IHC-P and IF listings for human samples, with IHC and IF images from human heart tissue (A05019 catalog; A05019 image captions).
Real IHC data
Anti-Homeodomain-only protein HOPX Antibody
Cat # A05019
A05019 is listed for human IHC-P and IF (A05019 catalog: applications, reactivity). Its own images show human heart tissue stained by IHC at 5 μg/mL and IF at 20 μg/mL (A05019 IHC and IF image captions).
Which to pick: Choose A05019 for paraffin-section tissue IHC based on its IHC-P listing; its own IHC image shows human heart tissue at 5 μg/mL, but the fixative is unreported (A05019 catalog: applications; A05019 IHC image caption). Choose A05019 for tissue IF at 20 μg/mL; ICC validation is unreported (A05019 IF image caption; A05019 catalog: applications). No cross-species option is supported: A05019 lists Human reactivity, has a Rabbit host, and has no reported clonality (A05019 catalog: reactivity, host, clone).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q9BPY8 (HOP_HUMAN, Homeodomain-only protein).
- Human Protein Atlas. HOPX tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. HOPX subcellular location (ICC-IF): Mainly localized to the nuclear bodies. In addition localized to the cytosol..
- Human Protein Atlas. HOPX antibody validation summary (2 antibodies).
- Epigenetic silencing of HOPX is critically involved in aggressive phenotypes and patient prognosis in papillary thyroid cancer. Oncotarget 2019 — PMC6800262.
- A discrete subtype of neural progenitor crucial for cortical folding in the gyrencephalic mammalian brain. eLife 2020 — PMC7173966.
- Cancer specific promoter CpG Islands hypermethylation of HOP homeobox (HOPX) gene and its potential tumor suppressive role in pancreatic carcinogenesis. BMC cancer 2012 — PMC3488580.
- HOPX functions as a tumour suppressor in head and neck cancer. Scientific reports 2016 — PMC5146930.
- PubMed PMID:14516659 — UniProt-cited evidence.
- PubMed PMID:12573257 — UniProt-cited evidence.
- PubMed PMID:12759545 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for HOPX (Homeodomain-only protein). Expected localisation Nuclear and cytoplasmic staining (HPA tissue IHC). Antigen retrieval: Tris-EDTA pH 9.0 HIER, 95–98 °C, 20 min (rule: nuclear antigen). Validated IHC antibody: A05019 at 5 μg/mL (the product's own IHC validation image is shown). Controls (HPA): positive Duodenum (Endocrine cells); negative Adipose tissue. 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 A05019); 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-HOPX, 5 μg/mL (datasheet A05019); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A05019); Caveat: Staining varies by cell population within a tissue (HPA tissue IHC); Isoform / epitope: 4 isoforms; epitope coverage is unspecified (UniProt); Regulation: Down-regulated in some cancers (UniProt). Evidence: UniProt Q9BPY8, HPA tissue IHC · Enhanced, HPA ICC-IF · Mainly localized to the nuclear bodies. In addition localized to the…, Datasheet A05019, PMC6800262, PMC3488580. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/HOPX/immunohistochemistry-ihc-guide.