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HGD / Homogentisate 1,2-dioxygenase · IHC design guide
Design Immunohistochemistry for HGD
Plan HGD IHC in paraffin sections using cytoplasmic staining in hepatocytes and select glandular cells as a reference (HPA tissue IHC). This guide covers fixation consistency, controls, chromogenic detection, and cell-specific scoring (standard IHC practice).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
HGD 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 | Cytoplasmic in a subset of tissues (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic staining in hepatocytes and select glandular cells (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6 HIER, heat-mediated (datasheet A01909-1) | |
| Positive control | Endometrium+4 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); Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A01909-1) | |
| Caveat | Liver endogenous peroxidase may confound DAB staining (standard IHC practice) | |
| Regulation | Expression is enriched in kidney and liver (HPA RNA) | |
| Isoform / epitope | No annotated isoforms; one 1–445 chain with no signal peptide (UniProt) |
Section 1
Recommended HGD IHC & IF Protocols
The catalog antibody’s IHC-P protocol is paired with one published mouse tissue IHC protocol (PMC13200543).
| Sample | Paraffin-embedded human liver cancer tissue; fixative not specified (datasheet A01909-1) |
| Fixation | Image fixative and duration unreported (datasheet A01909-1); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat retrieval: Citrate pH 6, 20 min (datasheet A01909-1) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A01909-1) |
| Primary antibody | Rabbit anti-HGD, 0.5-1μg/ml recommended; image 2μg/ml (datasheet A01909-1) |
| Primary incubation | Overnight at 4 °C (datasheet A01909-1) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet A01909-1) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | HGD-positive staining in ciliated epithelial cells of endometrium (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in a subset of tissues including liver, kidney, gallbladder, gastrointestinal tract, prostate and thyroid gland. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated citrate retrieval at pH 6 (datasheet A01909-1); the published protocol does not specify retrieval (PMC13200543).
Section 2
What Is the Expected HGD Staining Pattern?
In paraffin section IHC, expect HGD staining in the cytoplasm of hepatocytes and selected epithelial cells, including gallbladder glandular cells and kidney proximal tubules (HPA tissue IHC: Enhanced). HPA reports high staining in hepatocytes and gallbladder glandular cells, and medium staining in proximal tubules (HPA tissue IHC). HGD has no annotated transmembrane segment (UniProt Q93099 topology).
| Hepatocytes show clear cytoplasmic staining; nearby cells may differ in intensity. | This fits the high hepatocyte signal reported by HPA (HPA tissue IHC: High in hepatocytes). Judge the pattern by the stained cell type and compartment, then compare intensity with other tissues under the same staining conditions (general IHC practice). |
| Gallbladder glandular cells stain strongly, while kidney proximal tubule cell bodies stain less strongly. | This matches HPA's high gallbladder and medium kidney observations (HPA tissue IHC). A medium result in proximal tubules can still be an expected positive; do not require it to match hepatocyte intensity (HPA tissue IHC: Medium in proximal tubules). |
| The dominant signal outlines membranes, fills nuclei, or lies outside cells. | That distribution conflicts with HPA's cytoplasmic tissue IHC profile (HPA tissue IHC). Treat it as a possible staining artifact or off-target signal and review morphology, control sections, and detection background before scoring it as HGD (general IHC practice). |
| Strong staining appears in adipocytes or bone marrow hematopoietic cells. | HPA reports HGD as not detected in those cell types (HPA tissue IHC: adipocytes and hematopoietic cells). Check whether the signal follows cells or deposits, and consider cross-reactivity or endogenous chromogen activity as possibilities (general IHC practice). |
| A liver control has no hepatocyte signal. | HPA reports high hepatocyte staining, making this a useful warning that the run may have failed (HPA tissue IHC: High in hepatocytes). Inspect section integrity and the staining workflow before interpreting absent signal in a test specimen (general IHC practice). |
💡Expected HGD appearanceCall positive IHC when hepatocyte or gallbladder glandular cell cytoplasm stains strongly, or kidney proximal tubule cell bodies stain moderately (HPA tissue IHC); dominant nuclear, membrane, or extracellular color warrants investigation (HPA tissue IHC: cytoplasmic profile; general IHC practice).
| Tissue and cell selection | HPA records high signal in hepatocytes and gallbladder glandular cells, medium signal in proximal tubules, and no detection in adipocytes or bone marrow hematopoietic cells (HPA tissue IHC). Select controls by the specific cell population examined. |
| IHC antibody validation | Both listed antibodies, HPA047374 and HPA052359, have Enhanced IHC status (HPA antibodies). The tissue profile also has Enhanced reliability, reflecting high agreement between antibody staining and RNA expression (HPA tissue IHC); this supports the pattern, not every individual slide. |
| Assay-dependent compartment readout | Tissue IHC is described as cytoplasmic, whereas ICC-IF has an approved Golgi apparatus location (HPA tissue IHC; HPA subcellular ICC-IF). Use the cytoplasmic tissue profile for paraffin section IHC; assess the Golgi observation in the separate IF/ICC context. |
| Protein topology and processing | HGD has no annotated transmembrane segment, signal peptide, propeptide, or processed chain beyond residues 1–445 (UniProt Q93099). These annotations provide no basis to expect a membrane rim or secreted deposit; they do not establish antigen retrieval or fixation sensitivity. |
| Situation | Likely cause | Next action |
|---|---|---|
| Liver hepatocytes are unstained. | The result conflicts with HPA's high hepatocyte signal (HPA tissue IHC); a failed staining step or unsuitable section is possible (general IHC practice). | Review section quality, antigen retrieval, primary antibody application, detection reagents, and a concurrently stained positive control (general IHC practice). Avoid calling the test specimen negative until the control works. |
| Only kidney shows weak or patchy color. | Proximal tubule cell bodies are a medium, rather than high, HPA tissue IHC population; other kidney cells should not be treated as equivalent controls (HPA tissue IHC). | Identify proximal tubules on the counterstained section and compare their cell-body signal with the run's liver control (HPA tissue IHC; general IHC practice). Score intensity in the specified cells. |
| Nuclei or cell borders dominate the stain. | The pattern differs from the cytoplasmic tissue IHC profile (HPA tissue IHC). Background or off-target binding is possible (general IHC practice). | Inspect a no-primary control, reagent distribution, and tissue morphology; adjust blocking, washing, or detection conditions if the control shows color (general IHC practice). |
| Adipocytes or bone marrow cells appear strongly positive. | HPA lists adipocytes and bone marrow hematopoietic cells as not detected (HPA tissue IHC). Cross-reactivity or endogenous detection activity is possible, especially if control sections also stain (general IHC practice). | Compare no-primary and positive controls, check whether color tracks cells, and evaluate endogenous enzyme blocking for chromogenic detection (general IHC practice). |
| Color is diffuse across cells and empty spaces. | This does not resemble HPA's cell-associated cytoplasmic tissue profile (HPA tissue IHC). Excess detection background or poorly cleared chromogen is possible (general IHC practice). | Check the no-primary control and inspect wash, blocking, and chromogen steps; interpret only cell-associated staining after background is resolved (general IHC practice). |
| An IF/ICC image looks Golgi-localized while IHC looks broadly cytoplasmic. | HPA reports approved Golgi localization in ICC-IF and cytoplasmic staining in tissue IHC (HPA subcellular ICC-IF; HPA tissue IHC). | Record the assay and specimen type with each observation. Apply the cytoplasmic criterion to paraffin section IHC and evaluate the ICC-IF observation under its separate guide (HPA tissue IHC; HPA subcellular ICC-IF). |
Experiment evidence
IHC & IF images showing HGD staining
A01909-1 · IHCIHC analysis of HGD using anti-HGD antibody (A01909-1). HGD was detected in paraffin-embedded section of human liver cancer tissue. Heat med
A01909-1 · IHCIHC analysis of HGD using anti-HGD antibody (A01909-1). HGD was detected in paraffin-embedded section of human prostatic cancer tissue. Heat
A01909-1 · IFIF analysis of HGD using anti-HGD antibody (A01909-1). HGD was detected in immunocytochemical section of CACO-2 cells. Enzyme antigen retrie
M01909-1 · IHCIHC analysis of HGD using anti-HGD antibody (M01909-1). HGD was detected in a paraffin-embedded section of human liver cancer tissue. Heat m
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000113924-HGD/tissue/cerebral+cortex#imid_12896735
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000113924-HGD/tissue/colon#imid_12896645
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000113924-HGD/tissue/liver#imid_12896646
HPA · GallbladderHuman Protein Atlas IHC, Gallbladder. Source: https://www.proteinatlas.org/ENSG00000113924-HGD/tissue/gallbladder#imid_12896622
HPA · ICC-IF Hep-G2Human Protein Atlas immunofluorescence, Hep-G2, antibody HPA052359; green = HGD, blue = nucleus, red = microtubules.
HPA · ICC-IF Hep-G2Human Protein Atlas immunofluorescence, Hep-G2, antibody HPA052359; green = HGD, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for HGD IHC & IF
🧪Run liver first: hepatocytes should stain (HPA: High in hepatocytes). Use adipose tissue as the negative: adipocytes should lack detectable staining (HPA: Not detected in adipocytes); on the liver slide, compare hepatocytes with morphologically identified nonhepatocyte cells, treating them as internal negatives only if they remain unstained (HPA: High in hepatocytes).
Positive control tissue: Endometrium (Ciliated epithelial cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show HGD in Hep-G2, RPTEC/TERT1, U2OS, with annotated localisation: Golgi apparatus (approved) (HPA subcellular).
Technical controls: Include a no-primary (secondary-only) slide, a matched nonimmune rabbit IgG control, and, where available, HGD-knockout tissue or a validated peptide-block control (A01909-1 tissue-IHC caption: rabbit primary; standard IHC practice). Quench endogenous peroxidase and check endogenous biotin background in liver because the reported detection uses a streptavidin–biotin complex with DAB (A01909-1 tissue-IHC caption: SABC/DAB).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected caption does not state the fixative (A01909-1 tissue-IHC caption). The reported paraffin-section staining used heat retrieval in citrate buffer at pH 6 for 20 minutes; this supports starting there but does not establish that retrieval is required for every specimen (A01909-1 tissue-IHC caption). The supplied evidence does not establish that frozen sections or IF are easier; liver endogenous biotin may complicate the reported SABC/DAB detection (A01909-1 tissue-IHC caption; standard IHC practice).
HPA tissue IHC evidence for HGD
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 |
|---|---|---|---|---|
| Endometrium | Ciliated epithelial cells | High | Protein (IHC) | HPA → |
| Gallbladder | Glandular cells | High | Protein (IHC) | HPA → |
| Liver | Hepatocytes | High | Protein (IHC) | HPA → |
| Duodenum | Glandular cells | Medium | Protein (IHC) | HPA → |
| Kidney | Proximal tubules (cell body) | 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 → |
| Bone marrow | Hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Breast | Adipocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced HGD IHC Tips
Troubleshoot HGD staining in paraffin sections using the catalog antibody’s tissue image, tissue expression data, and matched controls.
Where should I start if HGD staining is weak after antigen retrieval?
Start with heat-mediated citrate buffer at pH 6 for 20 minutes (datasheet A01909-1; caption A01909-1). The catalog antibody’s paraffin-section image used that retrieval before overnight incubation at 4°C with 2 μg/mL primary antibody (caption A01909-1). Check a liver section for hepatocyte staining alongside the problem section, because hepatocytes show high HGD staining in the tissue survey (HPA tissue IHC). If both are weak, compare controlled retrieval times and primary-antibody concentrations while keeping detection and development matched; excessive heating can damage tissue morphology (standard IHC practice).
Could fixation explain inconsistent HGD staining across paraffin sections?
HGD-specific sensitivity to fixation is unknown from the supplied evidence; the catalog image identifies a paraffin section but does not state its fixative (caption A01909-1). Record each specimen’s fixative and processing history, then compare sections processed together before attributing a staining difference to HGD expression (standard IHC practice). Underfixation can impair morphology, while prolonged fixation can reduce antibody access to epitopes; neither effect has been demonstrated specifically for HGD here (standard IHC practice). Keep retrieval at citrate pH 6 for 20 minutes during the comparison, and include a concurrently processed positive control (caption A01909-1; standard IHC practice).
How should I assess cytoplasmic and Golgi-like HGD staining in tissue?
Judge the chromogenic result first against cytoplasmic staining in the expected tissue cells; the tissue survey describes that pattern across several organs (HPA tissue IHC). Its cell-image survey places HGD at the Golgi apparatus, whereas UniProt supplies no subcellular annotation, so a Golgi-like accent within cytoplasm is plausible but cannot be required in every tissue section (HPA subcellular; UniProt Q93099). HGD has no annotated transmembrane segment, making a crisp cell-surface-only pattern a reason to investigate specificity (UniProt Q93099 topology). Compare hepatocytes and kidney proximal tubules with nearby cells at the same DAB development time, and review morphology before assigning compartment-specific scores (HPA tissue IHC; standard IHC practice).
What if an HGD antibody stains some cells but misses others?
The supplied record lists one HGD chain spanning residues 1–445 and no annotated isoforms, so isoform switching has no support as an explanation here (UniProt Q93099). The catalog antibody’s epitope is unspecified in the supplied image evidence; do not assume that its binding region includes acetyllysine 98 or succinyllysine 414 (caption A01909-1; UniProt Q93099). Those modifications warrant consideration only if epitope mapping later places binding near them, rather than serving as a diagnosis of weak staining (UniProt Q93099; standard IHC practice). Compare matched retrieval and antibody conditions in an expected positive section, then investigate cell composition and section quality before proposing an epitope-specific effect (HPA tissue IHC; standard IHC practice).
How can IF help assess an ambiguous HGD IHC pattern?
Use IF as a separate localisation check, pairing HGD with a marker that identifies the expected cell population, such as hepatocytes in liver sections (HPA tissue IHC; standard IF practice). HGD has no annotated transmembrane segment, and cell-image evidence places it at the Golgi; allow antibody access to intracellular epitopes with appropriate permeabilisation after fixation (UniProt Q93099 topology; HPA subcellular; standard IF practice). Select fluorophores in channels with low measured tissue autofluorescence and include single-stain and no-primary controls before interpreting overlap (standard IF practice). Compare any punctate Golgi signal with the broader cytoplasmic pattern reported in tissue IHC, without treating the two imaging methods as identical (HPA subcellular; HPA tissue IHC).
How do I reduce diffuse DAB background while preserving HGD signal?
First inspect a no-primary control and a positive liver section developed together; diffuse color in both points toward detection or tissue background (HPA tissue IHC; standard IHC practice). The catalog image used 10% goat serum, 2 μg/mL primary antibody overnight at 4°C, and a biotin-based DAB detection system (caption A01909-1). Check peroxidase blocking, secondary-only staining, washing, and DAB development time before lowering primary concentration; these are general chromogenic IHC controls (standard IHC practice). Because the illustrated system uses biotin detection, assess a no-primary control for endogenous biotin contribution where background persists (caption A01909-1; standard IHC practice).
How should I score HGD staining across heterogeneous tissue sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the eligible cell population before scoring: hepatocytes show high staining, whereas kidney proximal tubules show medium staining in the tissue survey (HPA tissue IHC). For each region, record the percentage of viable eligible cells at intensity 0, 1+, 2+, and 3+, then calculate an H-score from 0–300 (standard IHC practice). Normalize positive-cell counts to all eligible cells, or positive area to the measured viable tissue area, while holding imaging and DAB thresholds constant (standard IHC practice). Exclude folds, damaged edges, and necrotic areas, and report regions separately when their cell composition differs (standard IHC practice).
Which staining features argue against a true HGD-positive result?
A convincing result follows the expected cell distribution and cytoplasmic tissue pattern, including hepatocytes and kidney proximal tubules, with appropriate negative controls (HPA tissue IHC; standard IHC practice). Strong membrane-only or nuclear-only staining deserves scrutiny because HGD has no annotated transmembrane segment and the cell-image survey places it at the Golgi (UniProt Q93099 topology; HPA subcellular). Staining confined to section edges, necrotic regions, or a no-primary control can reflect processing or detection artefacts (standard IHC practice). Review peroxidase blocking and the biotin-based detection controls before calling unusual DAB deposits HGD expression (caption A01909-1; standard IHC practice).
Boster reagents
Best HGD / Homogentisate 1,2-dioxygenase IHC Antibodies
A01909-1 and M01909-1 each have a human liver cancer paraffin-section IHC image (catalog IHC captions); A01909-1 also has a CACO-2 cell IF image (A01909-1 IF caption).
Real IHC data
Anti-HGD Antibody ®
Cat # A01909-1
Real IHC data
Anti-HGD Antibody ® (monoclonal, 2F11E1)
Cat # M01909-1
A01909-1 lists IHC, IF, and ICC applications and human, mouse, and rat reactivity (catalog applications/reactivity); its images show human liver cancer paraffin-section IHC and CACO-2 cell IF (A01909-1 image captions). M01909-1 lists human reactivity and IHC, but no IF application (catalog applications/reactivity); its image shows human liver cancer paraffin-section IHC (M01909-1 IHC caption).
Which to pick: For human tissue IHC, M01909-1 offers a monoclonal option (catalog: clone 2F11E1); its paraffin-section example used EDTA pH 8 retrieval, and the fixative is unreported (M01909-1 IHC caption). For IF/ICC or mouse and rat reactivity, choose A01909-1 (catalog applications/reactivity); its IF image uses CACO-2 cells (A01909-1 IF caption). Its tissue IHC example used paraffin sections and citrate pH 6 retrieval, with fixative unreported (A01909-1 IHC caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q93099 (HGD_HUMAN, Homogentisate 1,2-dioxygenase).
- Human Protein Atlas. HGD tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. HGD subcellular location (ICC-IF): Localized to the Golgi apparatus..
- Human Protein Atlas. HGD antibody validation summary (2 antibodies).
- Demarcated redness associated with increased vascular density/size: a useful marker of flat-type dysplasia in patients with ulcerative colitis. Endoscopy international open 2021 — PMC8041573.
- Specific knockout of kidney homogentisate 1,2-dioxygenase reveals that local metabolism of tyrosine and homogentisic acid is negligible in alkaptonuria. Human molecular genetics 2026 — PMC13200543.
- Expression analysis of Barrett's esophagus-associated high-grade dysplasia in laser capture microdissected archival tissue. Clinical cancer research : an official journal of the American Association for Cancer Research 2008 — PMC2701739.
- Expression of ATF6 as a marker of pre-cancerous atypical change in ulcerative colitis-associated colorectal cancer: a potential role in the management of dysplasia. Journal of gastroenterology 2018 — PMC5910497.
- PubMed PMID:8782815 — UniProt-cited evidence.
- PubMed PMID:9244427 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for HGD (Homogentisate 1,2-dioxygenase). Expected localisation Cytoplasmic in a subset of tissues (HPA tissue IHC). Antigen retrieval: Citrate pH 6 HIER, heat-mediated (datasheet A01909-1). Validated IHC antibody: A01909-1 at 0.5-1μg/ml recommended; image 2μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Endometrium (Ciliated epithelial cells); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human liver cancer tissue; fixative not specified (datasheet A01909-1); 2. Fixation: Image fixative and duration unreported (datasheet A01909-1); verify before use.; 3. Sectioning: 4–5 µm sections on charged slides (standard); 4. Deparaffinisation: Xylene, graded ethanol series to water (standard); 5. Antigen retrieval: Heat retrieval: Citrate pH 6 , 20 min (datasheet A01909-1); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A01909-1); 8. Primary antibody: Rabbit anti-HGD, 0.5-1μg/ml recommended; image 2μg/ml (datasheet A01909-1); 9. Primary incubation: Overnight at 4 °C (datasheet A01909-1). Watch for: Fixation: Keep fixation consistent across paraffin 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 A01909-1); Caveat: Liver endogenous peroxidase may confound DAB staining (standard IHC practice); Isoform / epitope: No annotated isoforms; one 1–445 chain with no signal peptide (UniProt); Regulation: Expression is enriched in kidney and liver (HPA RNA). Evidence: UniProt Q93099, HPA tissue IHC · Enhanced, HPA ICC-IF · Golgi apparatus., Datasheet A01909-1, PMC13200543. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/HGD/immunohistochemistry-ihc-guide.