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GCNT3 / Beta-1,3-galactosyl-O-glycosyl-glycoprotein beta-1,6-N-acetylglucosaminyltransferase 3 · IHC design guide
Design Immunohistochemistry for GCNT3
This GCNT3 paraffin-section IHC guide uses the granular cytoplasmic pattern in gastrointestinal mucosa to set staining expectations (HPA tissue IHC). It highlights the Golgi membrane location (UniProt) and cautions that presumed off-target binding can complicate interpretation (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
GCNT3 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 | Granular cytoplasm in gastrointestinal mucosa (HPA tissue IHC) | |
| Staining pattern | Granular cytoplasmic staining in gastrointestinal mucosa (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Appendix+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 A08909-1) | |
| Caveat | Presumed off-target binding may mimic GCNT3 staining (HPA tissue IHC) | |
| Regulation | Expression regulation is not annotated (UniProt) | |
| Isoform / epitope | No isoforms; lumenal vs cytoplasmic epitope affects access (UniProt) |
Section 1
Recommended GCNT3 IHC & IF Protocols
The catalog antibody protocol is accompanied by 3 published GCNT3 IHC protocols (PMC11651748; PMC13376619; PMC13126822).
| Sample | Paraffin-embedded human lung cancer tissue; fixative not specified (datasheet A08909-1) |
| Fixation | Image fixative and duration unreported (datasheet A08909-1); 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 citrate buffer, pH 6.0, 20 min at 95–98 °C (standard rule: cytoplasmic / membrane 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-GCNT3, 1:100-1:300 (datasheet A08909-1) |
| 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 | GCNT3-positive staining in enterocytes of appendix (HPA tissue IHC: High). HPA tissue profile: Granular cytoplasmic expression in gastrointestinal mucosa. No signal in the no-primary control. |
💡Decision noteStart with citrate pH 6.0 retrieval at 95–98 °C for 20 min (this page’s antigen-retrieval rule); compare the published conditions if staining needs optimization.
Section 2
What Is the Expected GCNT3 Staining Pattern?
GCNT3 is a single-pass Golgi membrane protein with a short cytoplasmic tail and a luminal catalytic region (UniProt O95395 topology). In paraffin-section IHC, expect granular cytoplasmic staining chiefly in gastrointestinal mucosal cells, including appendix and rectal enterocytes (HPA: tissue IHC). HPA rates the tissue profile Enhanced, while noting medium agreement with RNA data and presumed off-target staining that it disregarded (HPA: reliability description).
| Granular cytoplasmic staining in appendix or rectal enterocytes, with a clear cell outline. | This fits the reported gastrointestinal mucosal pattern and the High staining levels in these cells (HPA: tissue IHC). A Golgi-associated target can appear as cytoplasmic granules in chromogenic sections; the stain alone does not resolve the Golgi membrane (UniProt O95395 localization; general IHC practice). |
| Predominantly nuclear staining, or a continuous outline of the cell surface. | Neither is the expected dominant compartment for GCNT3, which is assigned to the Golgi membrane and has one membrane-spanning segment (UniProt O95395 localization and topology). Treat an isolated pattern of this kind as suspect; compare it with a positive tissue control and a detection-only control before assigning it to GCNT3 (general IHC practice). |
| Strong staining in cells reported as Not detected, such as adipocytes in adipose tissue or bronchial respiratory epithelial cells. | These cells are poor matches for the HPA tissue profile (HPA: adipose tissue and bronchus IHC). Consider antibody cross-reactivity or endogenous chromogen-generating activity, and inspect detection controls (general IHC practice). HPA reports that presumed off-target binding was observed and disregarded when assessing its tissue pattern (HPA: reliability description). |
| Uniform colour across epithelium, connective tissue and empty areas, without discernible cell-associated granules. | This does not resemble the reported granular cytoplasmic expression in gastrointestinal mucosa (HPA: tissue IHC profile). Broad, structure-independent colour can arise from background in chromogenic IHC; compare a section processed without primary antibody and assess blocking, washing and chromogen development (general IHC practice). |
| No staining in appendix or rectal enterocytes while the tissue morphology remains readable. | A negative result conflicts with the High levels reported for those cells (HPA: appendix and rectum IHC). First verify that the positive-control section and detection reagents worked; then review antibody dilution and antigen-retrieval conditions as general IHC variables (general IHC practice). The supplied sources do not establish GCNT3-specific fixation sensitivity. |
💡Expected GCNT3 appearanceCall a positive result when appendix or rectal enterocytes show convincing granular cytoplasmic staining, consistent with their High HPA levels and Golgi localization; dominant nuclear, continuous cell-surface or structure-independent colour is suspect (HPA: tissue IHC; UniProt O95395 localization; general IHC practice).
| Cell and tissue selection | Appendix and rectal enterocytes are reported High; colon enterocytes and stomach glandular cells are Medium (HPA: tissue IHC). Salivary and gallbladder glandular cells are Low (HPA: tissue IHC). Interpret a weak section against its stated cell type rather than expecting equal intensity across organs. |
| Compartment and epitope orientation | GCNT3 spans the membrane at residues 7–26; residues 1–6 are cytoplasmic and 27–438 are luminal (UniProt O95395 topology). These boundaries support a Golgi-associated interpretation but do not identify the antibody epitope or establish a GCNT3-specific retrieval requirement. |
| Evidence strength | HPA calls tissue IHC Enhanced and lists HPA011154 as IHC Enhanced, but describes medium staining-to-RNA consistency and disregarded presumed off-target binding (HPA: tissue reliability; HPA: antibody validation). An unexpected positive needs control-based review even when the expected pattern is present elsewhere. |
| IF/ICC Q&A: what localization is expected? | Mainly Golgi, with additional vesicle localization; HPA lists ICC as Approved for HPA011154 and images from A-549 and Hep-G2 (HPA: subcellular ICC-IF; HPA: antibody validation). This localization check belongs to IF/ICC interpretation and supplies no paraffin-section IF protocol. |
| Situation | Likely cause | Next action |
|---|---|---|
| The expected appendix or rectum control is blank. | Possible failure of primary staining, detection or a general IHC preparation step; these enterocytes are reported High (HPA: tissue IHC). | Check a known-working positive section and reagent run, then review antibody dilution and retrieval against the antibody's validated IHC instructions (general IHC practice). Do not infer a GCNT3-specific fixation effect from this result. |
| Granules are visible, but most of the section is also coloured. | The broad signal may be detection background rather than the cell-associated granular pattern reported by HPA (HPA: tissue IHC; general IHC practice). | Compare the primary-omission control; review blocking, washes and chromogen development before scoring cell-specific staining (general IHC practice). |
| Nuclei dominate the staining. | A nuclear-dominant result disagrees with the Golgi assignment (UniProt O95395 localization; HPA: subcellular ICC-IF). | Check the positive tissue and primary-omission controls, then reassess whether cytoplasmic granules are independently discernible (general IHC practice). |
| An unexpected cell population stains strongly. | Cross-reactivity or endogenous detection activity is possible (general IHC practice); HPA notes disregarded presumed off-target binding (HPA: reliability description). | Identify the stained cell type, compare its HPA tissue entry, and inspect detection controls before reporting GCNT3 positivity (HPA: tissue IHC; general IHC practice). |
| A Low tissue stains more weakly than the positive control. | Salivary and gallbladder glandular cells are reported Low, whereas appendix and rectal enterocytes are High (HPA: tissue IHC). | Score intensity within the relevant cell population and compare with the same-run positive and detection controls; do not require equal intensity across tissues (HPA: tissue IHC; general IHC practice). |
| Kidney staining is present, but its location seems unexpected. | HPA reports High staining in Bowman's capsule; it does not describe all kidney cells as High (HPA: kidney IHC). | Locate the stained structure on the counterstained section before scoring, and distinguish Bowman's capsule staining from broad renal background (HPA: kidney IHC; general IHC practice). |
Experiment evidence
IHC & IF images showing GCNT3 staining
A08909-1 · IHCImmunohistochemistry validation of GCNT3 using Anti-GCNT3 Antibody (A08909-1). Immunohistochemical analysis of paraffin-embedded human lung
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000140297-GCNT3/tissue/cerebral+cortex#imid_3058424
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000140297-GCNT3/tissue/colon#imid_3058342
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000140297-GCNT3/tissue/liver#imid_3058343
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000140297-GCNT3/tissue/kidney#imid_3058344
HPA · ICC-IF A-549Human Protein Atlas immunofluorescence, A-549, antibody HPA011154; green = GCNT3, blue = nucleus, red = microtubules.
HPA · ICC-IF A-549Human Protein Atlas immunofluorescence, A-549, antibody HPA011154; green = GCNT3, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for GCNT3 IHC & IF
🧪Run rectum first: enterocytes should stain (HPA: High in rectal enterocytes). Use adipose tissue as the negative tissue (HPA: Not detected in adipocytes); adjacent stromal cells on the rectal slide should show background-level chromogen as provisional internal negatives, without assuming they lack GCNT3.
Positive control tissue: Appendix (Enterocytes, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show GCNT3 in A-549, Hep-G2, with annotated localisation: Golgi apparatus (approved) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control and a host- and subclass-matched isotype control for a monoclonal primary, or species-matched nonimmune IgG for a polyclonal primary. Use a GCNT3-knockout sample or immunizing-peptide competition if available; in rectal sections, quench endogenous peroxidase, block endogenous biotin if using avidin–biotin detection, and assess luminal background.
⚠️Feasibility: No target-specific fixation window or fixation effect is reported; the selected A08909-1 paraffin-section caption gives a 1:100 dilution but does not state the fixative (selected-SKU tissue-IHC caption). Retrieval dependency is unreported, and the supplied evidence does not establish whether frozen sections or IF/ICC are easier; ICC-IF images exist for A-549 and Hep-G2 (HPA subcellular). In rectal sections, assess luminal mucus-associated background separately from intracellular staining; GCNT3 is mainly Golgi-localized, with additional vesicular localization (HPA subcellular).
HPA tissue IHC evidence for GCNT3
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Enhanced — Medium consistency between antibody staining and RNA expression data. Presumed off target binding observed and disregarded.). 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 |
|---|---|---|---|---|
| Appendix | Enterocytes | High | Protein (IHC) | HPA → |
| Kidney | Bowman's capsule | High | Protein (IHC) | HPA → |
| Rectum | Enterocytes | High | Protein (IHC) | HPA → |
| Colon | Enterocytes | Medium | Protein (IHC) | HPA → |
| Duodenum | Endocrine cells | 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 GCNT3 IHC Tips
Troubleshoot GCNT3 staining in paraffin sections by checking retrieval, cell type and Golgi localisation before comparing signal intensity (UniProt O95395; HPA tissue IHC).
What should I change when GCNT3 staining is weak after retrieval?
Start with citrate buffer at pH 6.0, heated to 95–98 °C for 20 min (page retrieval setting). If signal remains weak, compare a longer retrieval time on adjacent sections while checking for tissue damage (standard IHC practice). Keep the chromogenic detection steps and antibody dilution constant during that comparison so a stronger signal can be attributed to retrieval (standard IHC practice). The selected paraffin-section caption reports antibody dilution at 1:100 but does not state a retrieval method (A08909-1 caption). Score staining in expected epithelial cells and its granular cytoplasmic pattern, rather than treating diffuse colour as improvement (HPA tissue IHC).
How can I assess whether fixation is causing weak GCNT3 staining?
Target-specific sensitivity to fixation is unknown from the supplied evidence; the selected caption identifies paraffin-embedded tissue but does not state its fixative (A08909-1 caption). Record fixative, fixation duration and processing history for each block before comparing staining (standard IHC practice). Compare sections of similar thickness from consistently processed blocks using the same pH 6.0, 20 min retrieval setting (page retrieval setting; standard IHC practice). Include an expected positive cell population on each run, such as appendix enterocytes, and evaluate whether its staining also falls (HPA: High in appendix enterocytes; standard IHC practice). A difference between blocks warrants a controlled processing comparison, not a claim of proven GCNT3 fixation sensitivity (standard IHC practice).
Where should GCNT3 staining appear in a convincing paraffin section?
Expect a granular cytoplasmic pattern in gastrointestinal mucosa, consistent with GCNT3 being mainly Golgi-localised (HPA tissue IHC; HPA subcellular; UniProt O95395). The protein has an N-terminal membrane anchor at residues 7–26, with residues 27–438 on the lumenal side (UniProt O95395 topology). Use appendix or rectum enterocytes as pattern references, since both are reported High in tissue IHC (HPA: High in appendix enterocytes and rectum enterocytes). Check that chromogen is concentrated within the relevant cells rather than uniformly coating the tissue or collecting at section edges (standard IHC practice). Predominantly nuclear staining does not match the reported Golgi localisation and should prompt a specificity check (HPA subcellular; standard IHC practice).
Could an epitope mismatch explain different GCNT3 staining patterns?
The supplied record lists 0 isoforms, so these data do not support an isoform-specific explanation for differing stains (UniProt O95395). GCNT3 has a short cytoplasmic segment at residues 1–6, a transmembrane segment at 7–26 and a lumenal region at 27–438 (UniProt O95395 topology). Check the antibody's stated immunogen or epitope before comparing reagents, because its location is not supplied here (standard IHC practice; supplied antibody evidence). A glycosylation site is annotated at residue 289, but its effect on this antibody's staining has not been established (UniProt O95395; supplied antibody evidence). If two reagents disagree, compare their expected-cell and Golgi patterns on adjacent sections under matched retrieval and detection conditions (HPA tissue IHC; HPA subcellular; standard IHC practice).
How should I cross-check a GCNT3 IHC pattern with multiplex immunofluorescence?
Use IF as an independent localisation check: HPA reports GCNT3 mainly in the Golgi apparatus, with additional vesicular localisation (HPA subcellular). Pair GCNT3 with a marker identifying the epithelial cell population under study, then verify that the two signals occupy the expected cells (HPA tissue IHC; standard IF practice). Select spectrally separated fluorophores and consider a far-red channel for GCNT3 if the tissue has substantial shorter-wavelength autofluorescence (standard IF practice). If the antibody epitope lies in residues 27–438, optimise permeabilisation for access to the lumenal region; its actual epitope is unspecified (UniProt O95395 topology; supplied antibody evidence; standard IF practice). Validate IF fixation and permeabilisation independently, since the supplied paraffin-section caption gives neither an IF method nor fixation details (A08909-1 caption).
How do I reduce diffuse brown background without losing GCNT3 signal?
First separate granular intracellular signal from diffuse deposit by examining expected enterocytes alongside a no-primary control (HPA tissue IHC; standard IHC practice). Block endogenous peroxidase before chromogenic detection and use an appropriate protein block, following the detection system's instructions (standard IHC practice). If background persists, compare a lower primary-antibody concentration and shorter chromogen development while keeping retrieval at pH 6.0 for 20 min (page retrieval setting; standard IHC practice). The selected caption reports 1:100 antibody dilution in paraffin-embedded human lung cancer, which is a reference point rather than a universal optimum (A08909-1 caption). HPA notes presumed off-target binding in its tissue assessment, so corroborate any new staining pattern with controls (HPA tissue IHC).
How should I quantify GCNT3 staining across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and scoring region before reading the slides; HPA reports High staining in appendix and rectum enterocytes (HPA tissue IHC). For chromogenic staining, record the percentage of positive target cells and an intensity category, then calculate an H-score from 0–300 if appropriate (standard IHC practice). Normalise to the number of evaluable target cells or tissue area, rather than total section area when compartments differ (standard IHC practice). Apply identical retrieval at pH 6.0 for 20 min, imaging settings and scoring thresholds across compared sections (page retrieval setting; standard IHC practice). Exclude folds, edges and necrotic regions from analysis, and document exclusions before comparing groups (standard IHC practice).
When should I doubt an apparently positive GCNT3 IHC result?
Treat granular cytoplasmic staining in the expected epithelial population as more consistent with the reported pattern than isolated nuclear colour (HPA tissue IHC; HPA subcellular). GCNT3 is described as a Golgi membrane enzyme, and HPA reports High staining in appendix and rectum enterocytes (UniProt O95395; HPA tissue IHC). Re-examine staining limited to tissue edges, folds or necrotic areas, since those patterns can arise from processing and detection artefacts (standard IHC practice). Check a no-primary control and the peroxidase-blocking step when brown deposit appears broadly across cell types (standard IHC practice). HPA rates tissue staining Enhanced while noting medium RNA agreement and disregarded presumed off-target binding, so interpret unexpected cell populations cautiously (HPA tissue IHC).
Boster reagents
Best GCNT3 / Beta-1,3-galactosyl-O-glycosyl-glycoprotein beta-1,6-N-acetylglucosaminyltransferase 3 IHC Antibodies
The catalog antibody has a paraffin-section IHC image from human lung cancer (image caption); IF is listed, but no IF image is supplied (catalog applications; image payload).
Real IHC data
Anti-GCNT3 Antibody
Cat # A08909-1
A08909-1 will render with an IHC image of paraffin-embedded human lung cancer at 1:100 (image caption). A08909-1 lists IHC and IF applications and Human, Mouse and Rat reactivity; no IF image is supplied (catalog applications and reactivity; image payload).
Which to pick: Choose A08909-1 for paraffin-section tissue IHC because its own image caption documents that preparation; the fixative is unreported (image caption). For IF, A08909-1 lists a 1:200–1:1000 dilution, but ICC is not separately listed and no IF image is supplied (catalog applications and dilution; image payload). A08909-1 also lists Human, Mouse and Rat reactivity, while clonality is unreported (catalog reactivity and clone field).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry O95395 (GCNT3_HUMAN, Beta-1,3-galactosyl-O-glycosyl-glycoprotein beta-1,6-N-acetylglucosaminyltransferase 3).
- Human Protein Atlas. GCNT3 tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. GCNT3 subcellular location (ICC-IF): Mainly localized to the Golgi apparatus. In addition localized to vesicles..
- Human Protein Atlas. GCNT3 antibody validation summary (1 antibodies).
- Global analysis of human glycosyltransferases reveals novel targets for pancreatic cancer pathogenesis. British journal of cancer 2020 — PMC7251111.
- GCNT3 promotes the proliferation, apoptosis, invasion, and migration of breast cancer through the PI3K/AKT pathway. Translational cancer research 2024 — PMC11651748.
- Identification of GCNT3 as a glycometabolism-associated biomarker in endometrial cancer. Scientific reports 2026 — PMC13376619.
- GCNT3 and ST3GAL1 expression correlates with HER2 status and MUC1/β-catenin/Cyclin D1 axis in breast cancer. BMC cancer 2026 — PMC13126822.
- PubMed PMID:9915862 — UniProt-cited evidence.
- PubMed PMID:9988682 — UniProt-cited evidence.
- PubMed PMID:17303715 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for GCNT3 (Beta-1,3-galactosyl-O-glycosyl-glycoprotein beta-1,6-N-acetylglucosaminyltransferase 3). Expected localisation Granular cytoplasm in gastrointestinal mucosa (HPA tissue IHC). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: A08909-1 at 1:100-1:300 (the product's own IHC validation image is shown). Controls (HPA): positive Appendix (Enterocytes); negative Adipose tissue. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Paraffin-embedded human lung cancer tissue; fixative not specified (datasheet A08909-1); 2. Fixation: Image fixative and duration unreported (datasheet A08909-1); 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 citrate buffer, pH 6.0 , 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-GCNT3, 1:100-1:300 (datasheet A08909-1); 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 A08909-1); Caveat: Presumed off-target binding may mimic GCNT3 staining (HPA tissue IHC); Isoform / epitope: No isoforms; lumenal vs cytoplasmic epitope affects access (UniProt); Regulation: Expression regulation is not annotated (UniProt). Evidence: UniProt O95395, HPA tissue IHC · Enhanced, HPA ICC-IF · Mainly localized to the Golgi apparatus. In addition localized to…, Datasheet A08909-1, PMC11651748, PMC13376619. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/GCNT3/immunohistochemistry-ihc-guide.