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ARFGAP3 / ADP-ribosylation factor GTPase-activating protein 3 · IHC design guide
Design Immunohistochemistry for ARFGAP3
Plan chromogenic paraffin-section IHC for ARFGAP3 with an antibody dilution of 1:50–1:200 (datasheet: A06839-1). Assess cytoplasmic staining in glandular cells while accounting for the low concordance between antibody staining and RNA expression (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
ARFGAP3 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 tissue (HPA tissue IHC); Golgi-associated expected (UniProt) | |
| Staining pattern | Cytoplasmic staining, most abundant in glandular cells (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Pancreas+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) | |
| Caveat | Antibody staining has low concordance with RNA levels (HPA tissue IHC) | |
| Regulation | Expression varies by tissue (UniProt) | |
| Isoform / epitope | Two isoforms; check splice-dependent epitope coverage (UniProt) |
Section 1
Recommended ARFGAP3 IHC & IF Protocols
The catalog antibody’s IHC-P protocol is accompanied by one published anterior vaginal wall IHC example (PMC7774809).
| Sample | Paraffin-embedded mouse stomach tissue; fixative not specified (datasheet A06839-1) |
| Fixation | Image fixative and duration unreported (datasheet A06839-1); 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-ARFGAP3, 1:50-1:200 (datasheet A06839-1) |
| 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 | ARFGAP3-positive staining in exocrine glandular cells of pancreas (HPA tissue IHC: Medium). HPA tissue profile: Cytoplasmic expression in several different tissue types, most abundant in glandular cells. No signal in the no-primary control. |
💡Decision noteStart with citrate pH 6.0 HIER at 95–98 °C for 20 min (page retrieval rule: cytoplasmic/membrane antigen).
Section 2
What Is the Expected ARFGAP3 Staining Pattern?
ARFGAP3 should appear in the cytoplasm of several tissue cell types, especially glandular cells (HPA: tissue IHC profile). A cytosolic pattern with possible Golgi enrichment is consistent with its reported locations (HPA: supported ICC-IF locations; UniProt Q9NP61: Golgi membrane and cytoplasm). ARFGAP3 has no transmembrane segment (UniProt Q9NP61: topology). Interpret tissue staining cautiously: HPA rates its IHC profile Approved but reports low consistency with RNA expression (HPA: tissue IHC reliability).
| Cytoplasmic staining in glandular cells of pancreas, prostate, salivary gland, rectum, stomach, or small intestine. | This fits the reported medium staining in those specified cell populations (HPA: tissue IHC). Assess the stained cell type within each section; the HPA result does not imply that every cell in a listed tissue is positive. |
| A concentrated perinuclear component accompanies broader cytoplasmic staining. | Golgi enrichment is plausible given the reported Golgi and cytoplasmic locations (UniProt Q9NP61: subcellular location; HPA: supported ICC-IF locations). In chromogenic IHC, morphology alone cannot prove that a stained structure is the Golgi. |
| Strong staining is confined to nuclei or another compartment inconsistent with the cytoplasm and Golgi. | Treat this as a suspect pattern and check controls and detection steps before calling it ARFGAP3: the supported locations are cytosol and Golgi (HPA: ICC-IF), while UniProt also notes peripheral punctate structures likely to be the ER–Golgi intermediate compartment (UniProt Q9NP61: subcellular location). |
| The main signal is in a cell population reported as undetected, such as adipocytes in adipose tissue. | Consider cross-reactivity or endogenous chromogenic activity, then verify the finding with appropriate controls (HPA: adipocytes not detected; general IHC practice). An HPA cell-level negative result should not be extended to every cell in that tissue. |
| The section shows widespread haze, or no signal appears in a selected medium-staining glandular population. | Haze limits assessment of the expected cellular pattern; absent signal in a selected positive population raises a technical or specimen-level question (HPA: tissue IHC; general IHC practice). Neither observation alone establishes ARFGAP3 abundance. |
💡Expected ARFGAP3 appearanceCall a section positive when glandular cells show discernible cytoplasmic staining, potentially with Golgi enrichment, at an intensity judged against controls; HPA reports medium staining in several glandular populations, whereas isolated nuclear signal or uniform haze is suspect (HPA: tissue IHC and supported ICC-IF locations; general IHC practice).
| Which cells provide a tissue benchmark? | HPA reports medium staining in pancreatic exocrine glandular cells and glandular cells of prostate, rectum, salivary gland, seminal vesicle, small intestine, and stomach; decidual cells in placenta are also medium (HPA: tissue IHC). These are observed patterns, not guarantees for every section. |
| How firm is the tissue-level expectation? | The antibody HPA000638 is Approved for IHC, while the tissue profile notes low consistency between antibody staining and RNA expression (HPA: antibody validation; HPA: tissue IHC reliability). Give controls and cell morphology weight when interpreting an unexpected result. |
| What does the protein record add to localisation? | UniProt places ARFGAP3 at the Golgi membrane and in cytoplasm, with possible peripheral puncta, and lists no transmembrane segment, signal peptide, propeptide, or cleavage into shorter chains (UniProt Q9NP61: location, topology, processing). Those annotations do not predict retrieval conditions or fixation sensitivity. |
| Could isoforms affect interpretation? | Two isoforms are listed (UniProt Q9NP61: isoforms). The supplied records do not identify which isoform the IHC antibody recognises, so an isoform-specific tissue or compartment prediction is unsupported. |
| IF/ICC: what pattern belongs on its separate guide page? | In ICC-IF, HPA supports a mainly cytosolic location with additional Golgi localisation and lists images from A-431, U-251MG, and U2OS; HPA000638 is Supported for ICC (HPA: subcellular; HPA: antibody validation). Those observations inform localisation, not an IHC protocol choice. |
| Situation | Likely cause | Next action |
|---|---|---|
| No stain in a selected medium-staining glandular population. | Possible loss of detectable signal during the IHC workflow or an uninformative section; HPA's medium rating is observational, not a guaranteed result (HPA: tissue IHC; general IHC practice). | Check a concurrently stained suitable positive section, tissue preservation, antibody incubation, retrieval conditions, and chromogenic detection controls (general IHC practice). Do not infer target-specific retrieval or fixation sensitivity from HPA. |
| Diffuse colour obscures cell boundaries throughout the section. | Nonspecific binding or excess detection background can obscure localisation (general IHC practice). | Review blocking, washing, primary antibody conditions, and a no-primary control; score only interpretable cellular staining after background is controlled (general IHC practice). |
| Colour persists in a no-primary control. | Endogenous detection activity or reagent-associated background may contribute to a chromogenic signal (general IHC practice). | Check the detection chemistry and its appropriate endogenous-activity controls before assigning the colour to ARFGAP3 (general IHC practice). |
| Staining is mainly nuclear. | The dominant compartment conflicts with the cytosolic and Golgi locations supported for ARFGAP3 (HPA: ICC-IF; UniProt Q9NP61: subcellular location). | Compare with a suitable positive section and no-primary control, and reassess morphology and background before scoring nuclear staining as target-specific (general IHC practice). |
| Unexpectedly strong signal appears in HPA-undetected adipocytes or cervical glandular cells. | Cross-reactivity or local detection background is possible; HPA lists those particular populations as not detected (HPA: tissue IHC; general IHC practice). | Verify the cell identity, compare controls, and repeat assessment with an independent validated antibody if available (general IHC practice). Avoid declaring the entire tissue negative from a cell-specific HPA entry. |
| A Golgi-like focus is absent, but cytoplasmic staining is clear. | Golgi signal may be difficult to distinguish from broader cytoplasmic colour in a chromogenic section (HPA: supported cytosolic and Golgi locations; general IHC practice). | Judge the result against tissue cell type, overall cytoplasmic pattern, and controls; reserve a precise Golgi assignment for localisation evidence suited to that question (general IHC practice). |
Experiment evidence
IHC & IF images showing ARFGAP3 staining
A06839-1 · IHCImmunohistochemistry of paraffin-embedded mouse stomach using ARFGAP3 antibody at dilution of 1:100 (x40 lens).
A06839-1 · IHCImmunohistochemistry of paraffin-embedded human liver cancer using ARFGAP3 antibody at dilution of 1:100 (x40 lens).
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000242247-ARFGAP3/tissue/cerebral+cortex#imid_226722
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000242247-ARFGAP3/tissue/colon#imid_226656
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000242247-ARFGAP3/tissue/liver#imid_226657
HPA · PancreasHuman Protein Atlas IHC, Pancreas. Source: https://www.proteinatlas.org/ENSG00000242247-ARFGAP3/tissue/pancreas#imid_226616
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA000638; green = ARFGAP3, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA000638; green = ARFGAP3, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for ARFGAP3 IHC & IF
🧪Run pancreas first and look for staining in exocrine glandular cells (Medium; HPA: pancreas). Run adipose tissue as the negative tissue, focusing on adipocytes (HPA: Not detected in adipocytes); on the pancreas slide, treat any unstained neighboring cells as an internal comparison, since the supplied HPA pancreas row does not identify a negative cell population.
Positive control tissue: Pancreas (Exocrine glandular cells, HPA Medium)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show ARFGAP3 in A-431, U-251MG, U2OS, with annotated localisation: Cytosol (supported) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control, an isotype control matched to the primary antibody’s host species, immunoglobulin class, and clonality, and a biological negative such as knockout tissue or peptide blocking when available (standard IHC practice). For chromogenic pancreas IHC, block endogenous peroxidase and check residual background on the no-primary slide (standard IHC practice).
⚠️Feasibility: A target-specific fixation window, fixation effect, and antigen-retrieval dependency are unreported in the supplied evidence; the selected 1:100 paraffin-section mouse stomach caption also leaves the fixative unreported (catalog antibody caption). The evidence does not establish whether frozen sections or IF are easier, although HPA reports ICC-IF localization mainly in the cytosol with additional Golgi localization (HPA: subcellular). In pancreas, exocrine granules can complicate interpretation of punctate staining, so compare the pattern with the negative controls (standard IHC practice; HPA: Medium in exocrine glandular cells).
HPA tissue IHC evidence for ARFGAP3
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Low 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 |
|---|---|---|---|---|
| Pancreas | Exocrine glandular cells | Medium | Protein (IHC) | HPA → |
| Placenta | Decidual cells | Medium | Protein (IHC) | HPA → |
| Prostate | Glandular cells | Medium | Protein (IHC) | HPA → |
| Rectum | Glandular cells | Medium | Protein (IHC) | HPA → |
| Salivary gland | Glandular cells | Medium | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Caudate | Glial cells | Not detected | Protein (IHC) | HPA → |
| Cerebellum | Cells in granular layer | Not detected | Protein (IHC) | HPA → |
| Cerebral cortex | Endothelial cells | Not detected | Protein (IHC) | HPA → |
| Cervix | Glandular cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced ARFGAP3 IHC Tips
Use compartment, cell type, and matched controls to troubleshoot ARFGAP3 staining in paraffin sections; treat the mouse stomach image as antibody-specific evidence.
What retrieval should I try first when ARFGAP3 staining is weak?
Start with citrate buffer at pH 6.0 for heat-induced retrieval at 95–98 °C for 20 minutes (page retrieval rule: cytoplasmic/membrane antigen). Allow sections to cool in the retrieval buffer, then compare staining with a matched section processed without retrieval (standard IHC practice). The catalog antibody has a paraffin-embedded mouse stomach image at 1:100, but its caption does not report retrieval conditions (caption: A06839-1). If staining remains weak, test EDTA at pH 9.0 on adjacent sections while keeping detection conditions constant (standard IHC optimisation). Judge improvement by clearer cytoplasmic staining in glandular cells without a comparable rise in background (HPA: cytoplasmic expression most abundant in glandular cells).
How should I assess whether fixation is masking ARFGAP3?
Target-specific fixation sensitivity is unknown: the selected mouse stomach caption identifies paraffin embedding but does not state a fixative (caption: A06839-1). Record the fixative and fixation duration for each specimen, then compare similarly processed sections before changing retrieval or antibody concentration (standard IHC practice). Overfixation can reduce epitope accessibility in paraffin IHC, but that general possibility does not establish an ARFGAP3-specific effect (standard IHC practice). If archival blocks differ in processing, include a consistently processed reference section in each run and interpret intensity differences cautiously (standard IHC practice). Do not infer fixation tolerance from tissue staining patterns or protein topology (HPA: tissue IHC profile; UniProt Q9NP61 topology).
Which staining pattern is plausible for ARFGAP3 in tissue sections?
Expect predominantly cytoplasmic signal, potentially concentrated near the Golgi, rather than an exclusively nuclear pattern (HPA: tissue IHC profile; HPA: supported cytosol and Golgi locations). ARFGAP3 is annotated at the Golgi membrane and in cytoplasm, with additional peripheral puncta likely corresponding to the ER–Golgi intermediate compartment (UniProt Q9NP61 subcellular location). Because it has no transmembrane segment, a continuous cell-surface rim is not its expected distribution (UniProt Q9NP61 topology). Assess signal within glandular cells and compare it with neighboring compartments on the same section (HPA: cytoplasmic staining most abundant in glandular cells; standard IHC practice). A hematoxylin counterstain helps distinguish perinuclear cytoplasm from nuclei (standard chromogenic IHC practice).
Could isoforms or modifications explain discordant ARFGAP3 staining?
ARFGAP3 has two annotated isoforms, but the supplied antibody caption does not identify its epitope or establish isoform selectivity (UniProt Q9NP61 isoforms; caption: A06839-1). Check the antibody's immunogen or epitope documentation before claiming that either isoform is absent from a stained cell (standard IHC validation practice). The annotated Arf-GAP domain spans residues 10–126, and several phosphoserines are recorded elsewhere in the sequence (UniProt Q9NP61 domains and modified residues). Those annotations alone cannot show whether fixation or retrieval affects this antibody's binding (UniProt Q9NP61 annotations; standard IHC interpretation). If isoform specificity matters, compare staining with an independently characterised epitope or an appropriate genetic control (standard IHC validation practice).
How can IF help check cell identity and intracellular pattern?
For a separate IF experiment, multiplex ARFGAP3 with a glandular epithelial marker such as pan-cytokeratin and assess overlap at the cell level (HPA: staining most abundant in glandular cells; standard IF practice). Select a spectrally separated far-red channel for a weak target signal when the tissue shows strong shorter-wavelength autofluorescence, and include unstained sections (standard IF practice). ARFGAP3 is cytoplasmic and Golgi-associated without a transmembrane segment, so use controlled permeabilisation to access intracellular epitopes while recognising that this antibody's epitope is unspecified (UniProt Q9NP61 localisation and topology; caption: A06839-1). Compare punctate or perinuclear signal with the broader cytosolic pattern, using single-label controls to assess bleed-through (HPA: supported cytosol and Golgi locations; standard IF practice).
What should I change when chromogenic staining is diffuse or widespread?
First compare a no-primary control and a matched section stained with the catalog antibody at the caption's 1:100 dilution (standard IHC practice; caption: A06839-1). Block endogenous peroxidase before HRP detection, and check whether background appears in the no-primary section after DAB development (standard chromogenic IHC practice). If background depends on the primary antibody, titrate it below 1:100 and strengthen protein blocking or washes while preserving the same retrieval conditions (standard IHC optimisation). Assess whether signal remains cytoplasmic in glandular cells rather than uniformly coating extracellular material (HPA: tissue IHC profile). Inspect section edges separately, since drying and uneven reagent coverage can produce misleadingly intense margins (standard IHC practice).
How should I quantify ARFGAP3 across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, using glandular-cell cytoplasm where that population is present (HPA: cytoplasmic expression most abundant in glandular cells; standard IHC practice). Report the percentage of positive cells and staining intensity, or calculate an H-score from intensity categories 0–3, giving a possible range of 0–300 (standard IHC scoring practice). Normalise counts to the number of eligible cells, or report positive-cell density per mm² of viable tissue when cell counting is impractical (standard IHC quantification practice). Use the same exposure, counterstain, region-selection rules, and reference section across batches (standard IHC practice). Keep cytoplasmic and nuclear scores separate because the supported localisation is cytosolic with an additional Golgi component (HPA: subcellular location).
When is apparent ARFGAP3 positivity likely to be an artefact?
Give greatest weight to reproducible cytoplasmic or perinuclear signal in appropriate cells, consistent with the supported cytosol and Golgi locations (HPA: subcellular location; standard IHC interpretation). Glandular-cell staining is plausible, while a strong isolated nuclear or cell-surface pattern warrants review of specificity and morphology (HPA: tissue IHC profile; UniProt Q9NP61 topology). Check whether staining concentrates at section edges or in necrotic areas, where tissue damage can distort chromogenic interpretation (standard IHC practice). Residual endogenous peroxidase is suspect if DAB signal persists in a no-primary control (standard chromogenic IHC practice). Finally, treat a visually convincing pattern as provisional because the tissue IHC record is Approved yet reports low consistency between antibody staining and RNA expression (HPA: reliability description).
Boster reagents
Best ARFGAP3 / ADP-ribosylation factor GTPase-activating protein 3 IHC Antibodies
A06839-1 has IHC images from paraffin-embedded mouse stomach and human liver cancer (catalog image captions). No IF/ICC data are listed (catalog: applications IHC, WB; IF images absent).
Real IHC data
Anti-ARFGAP3 Antibody
Cat # A06839-1
A06839-1 is listed for IHC and reacts with human and mouse (catalog: applications and reactivity). Its own IHC images show paraffin-embedded mouse stomach and human liver cancer at 1:100 (catalog image captions).
Which to pick: Choose A06839-1 for paraffin-section tissue IHC based on its own mouse stomach and human liver cancer images; its clonality is unspecified. No listed SKU has IF/ICC validation (catalog: A06839-1 applications IHC, WB; IF images absent). A06839-1 is the listed option for human and mouse IHC (catalog: reactivity Human, Mouse); its paraffin-section captions do not report a fixative (catalog image captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q9NP61 (ARFG3_HUMAN, ADP-ribosylation factor GTPase-activating protein 3).
- Human Protein Atlas. ARFGAP3 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. ARFGAP3 subcellular location (ICC-IF): Mainly localized to the cytosol. In addition localized to the Golgi apparatus..
- Human Protein Atlas. ARFGAP3 antibody validation summary (1 antibodies).
- Expression of ArfGAP3 in Vaginal Anterior Wall of Patients With Pelvic Floor Organ Prolapse in Pelvic Organ Prolapse and Non-Pelvic Organ Prolapse Patients. Female pelvic medicine & reconstructive surgery 2021 — PMC7774809.
- PubMed PMID:10704287 — UniProt-cited evidence.
- PubMed PMID:12529303 — UniProt-cited evidence.
- PubMed PMID:15461802 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for ARFGAP3 (ADP-ribosylation factor GTPase-activating protein 3). Expected localisation Cytoplasmic in tissue (HPA tissue IHC); Golgi-associated expected (UniProt). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: A06839-1 at 1:50-1:200 (the product's own IHC validation image is shown). Controls (HPA): positive Pancreas (Exocrine glandular cells); negative Adipose tissue. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Paraffin-embedded mouse stomach tissue; fixative not specified (datasheet A06839-1); 2. Fixation: Image fixative and duration unreported (datasheet A06839-1); 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-ARFGAP3, 1:50-1:200 (datasheet A06839-1); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across paraffin sections. (standard IHC practice; not target-specific); Caveat: Antibody staining has low concordance with RNA levels (HPA tissue IHC); Isoform / epitope: Two isoforms; check splice-dependent epitope coverage (UniProt); Regulation: Expression varies by tissue (UniProt). Evidence: UniProt Q9NP61, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the cytosol. In addition localized to the Golgi…, Datasheet A06839-1, PMC7774809. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/ARFGAP3/immunohistochemistry-ihc-guide.