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RAP1GAP / Rap1 GTPase-activating protein 1 · IHC design guide
Design Immunohistochemistry for RAP1GAP
Plan RAP1GAP chromogenic IHC in paraffin sections using the catalog antibody’s documented conditions (datasheet A03561-2). Compare staining with the cytoplasmic tissue pattern and cell-level controls reported by HPA (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
RAP1GAP 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 staining in most tissues (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic signal in most tissues, including brain (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A03561-2) | |
| Positive control | Adrenal gland+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 A03561-2) | |
| Caveat | Antibody staining and RNA expression have medium consistency (HPA tissue IHC) | |
| Regulation | Cortex expression exceeds spinal cord (UniProt) | |
| Isoform / epitope | Four isoforms; verify antibody epitope coverage (UniProt) |
Section 1
Recommended RAP1GAP IHC & IF Protocols
Compare the catalog antibody’s IHC-P protocol with 4 published RAP1GAP IHC protocols (datasheet: A03561-2; PMC12098257; PMC2822891; PMC5352382; PMC3531979).
| Sample | Paraffin-embedded human gastric adenocarcinoma tissue; fixative not specified (datasheet A03561-2) |
| Fixation | Image fixative and duration unreported (datasheet A03561-2); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat retrieval: EDTA pH 8.0 (datasheet A03561-2); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A03561-2) |
| Primary antibody | Rabbit anti-RAP1GAP, 2-5 μg/ml (datasheet A03561-2) |
| Primary incubation | Overnight at 4 °C (datasheet A03561-2) |
| Detection | HRP-conjugated secondary, DAB chromogen (datasheet A03561-2) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | RAP1GAP-positive staining in glandular cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in most tissues, including thryoid gland and brain. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA retrieval at pH 8.0 for the catalog antibody; a published gastric protocol used citrate at pH 6.0 (datasheet: A03561-2; PMC5352382).
Section 2
What Is the Expected RAP1GAP Staining Pattern?
In paraffin-section IHC, expect RAP1GAP mainly in the cytoplasm, including High staining in cerebral cortex neurons and selected glandular or epithelial cells (HPA tissue IHC). UniProt places RAP1GAP at the Golgi apparatus membrane and reports no transmembrane segment (UniProt P47736). HPA rates its tissue IHC reliability Enhanced, while noting medium consistency between antibody staining and RNA expression (HPA tissue IHC).
| Cerebral cortex neurons show clear cytoplasmic chromogen, with distinguishable cell outlines and limited background. | This fits a strong positive tissue and the reported cytoplasmic pattern (HPA: High in cerebral cortex neuronal cells; HPA tissue IHC profile). Score the neurons separately from surrounding cells; the tissue-level label does not assign every cell the same intensity. |
| Staining appears predominantly nuclear, with little or no cytoplasmic signal in otherwise positive cells. | A nuclear-only result conflicts with the reported IHC profile and approved ICC-IF cytosolic location (HPA tissue IHC; HPA subcellular). Treat it as a localisation concern and review controls, antibody specificity and image interpretation before assigning it to RAP1GAP. |
| Cells reported as undetected stain strongly, such as adipocytes beside other tissue elements. | HPA reports RAP1GAP as Not detected in adipocytes, specifically, rather than in every cell of adipose tissue (HPA: Not detected in adipocytes). Check cell identification and background controls; staining in an unexpected cell population may reflect cross-reactivity or endogenous chromogenic activity (general IHC practice). |
| Color is spread across tissue, including extracellular spaces, and cell boundaries are hard to resolve. | That distribution is less interpretable than cell-associated cytoplasmic staining (HPA tissue IHC profile). Uneven background can arise during blocking, washing or chromogenic detection (general IHC practice); compare a reagent control and review the entire section before scoring cells. |
| A cerebral cortex section has no detectable staining in neurons. | The absence conflicts with the reported High neuronal staining (HPA: cerebral cortex neuronal cells). It does not by itself establish biological absence. Check tissue identification, the IHC-validated antibody, antigen retrieval, primary-antibody application and detection controls (general IHC practice). |
💡Expected RAP1GAP appearanceCall a section positive when identifiable cerebral cortex neurons, or another HPA-listed positive cell population, show cell-associated cytoplasmic staining consistent with the reported High level; nuclear-only color or widespread cell-free deposit is suspect (HPA tissue IHC; general IHC practice).
| Tissue and cell selection | HPA reports High staining in cerebral cortex neurons, adrenal gland glandular cells and bronchial respiratory epithelial cells (HPA tissue IHC). It reports Not detected in skeletal-muscle myocytes and adipocytes (HPA tissue IHC). Choose and score the named cell population, since a tissue can contain several cell types. |
| Compartment interpretation | HPA describes cytoplasmic staining across most tissues and approved cytosolic localisation by ICC-IF; UniProt lists the Golgi apparatus membrane and no transmembrane segment (HPA tissue IHC; HPA subcellular; UniProt P47736). These records support a cytoplasmic call, but do not require a visibly Golgi-restricted chromogenic pattern. |
| IHC antibody evidence | HPA marks the tissue IHC assessment Enhanced and lists HPA001922 as IHC Enhanced; its summary notes medium agreement between staining and RNA data (HPA tissue IHC; HPA antibodies). That supports the published pattern while leaving an unexpected result to be checked with appropriate controls. |
| Isoforms and epitope coverage | UniProt lists 4 RAP1GAP isoforms (UniProt P47736). The supplied records give no antibody epitope or isoform coverage, so a difference between specimens cannot be attributed to a particular isoform from these data alone. |
| Processing and modifications | UniProt describes one chain spanning residues 1–663, no signal peptide, no propeptide and no listed glycosylation sites; it lists several phosphoserines (UniProt P47736). These annotations do not establish whether retrieval or fixation changes staining, because no target-specific sensitivity data are supplied. |
| IF/ICC Q: where should signal appear? | A: HPA approves cytosol as the main ICC-IF location and shows images for A-431 and U-251MG; CAB003851 has an Uncertain ICC assessment (HPA subcellular; HPA antibodies). Use the separate IF/ICC guide for that application; these data do not turn its images into an IHC protocol. |
| Situation | Likely cause | Next action |
|---|---|---|
| No signal in cortex neurons on a nominal positive section. | Possible tissue misidentification or an IHC workflow failure; HPA reports High neuronal staining there (HPA: cerebral cortex). | Confirm the neurons and section identity, then review retrieval, primary-antibody application, chromogen and a detection control in sequence (general IHC practice). |
| Only nuclei stain in cells expected to be positive. | The compartment differs from HPA's cytoplasmic IHC profile and approved cytosolic ICC-IF location (HPA tissue IHC; HPA subcellular). | Inspect the counterstain and no-primary control, then repeat with the IHC-validated antibody and score cytoplasmic signal separately (general IHC practice). |
| Adipocytes or skeletal-muscle myocytes stain strongly. | Those named cell populations are reported Not detected (HPA: adipocytes; HPA: skeletal-muscle myocytes). Cell misidentification, cross-reactivity or endogenous detection activity are possibilities (general IHC practice). | Recheck morphology and compare no-primary and detection-only controls; record staining by cell type rather than calling the whole tissue positive (general IHC practice). |
| Diffuse brown deposit obscures cellular localisation. | The slide cannot be compared reliably with HPA's cell-associated cytoplasmic profile (HPA tissue IHC). Blocking, washing or detection background may contribute (general IHC practice). | Review reagent controls and staining uniformity; optimise general blocking, washing and detection conditions before interpreting intensity (general IHC practice). |
| Staining differs between the catalog antibody and an HPA image. | HPA's Enhanced assessment describes its evaluated IHC evidence; the supplied records do not establish epitope equivalence or matching isoform coverage for the catalog antibody (HPA tissue IHC; UniProt P47736). | Verify the antibody's IHC-P validation and compare the same tissue, cell population and compartment before treating intensity differences as biological (general IHC practice). |
| A result changes after a retrieval or fixation adjustment. | Target-specific fixation effects are not established by the supplied assay evidence. Verify with a matched IHC source before attributing a result to fixation. | Keep processing and scoring consistent across compared sections, and use matched controls to determine whether the workflow change explains the result (general IHC practice). |
Experiment evidence
IHC & IF images showing RAP1GAP staining
A03561-2 · IHCIHC analysis of RAP1GAP using anti-RAP1GAP antibody (A03561-2). RAP1GAP was detected in a paraffin-embedded section of human gastric adenoca
A03561-2 · IHCIHC analysis of RAP1GAP using anti-RAP1GAP antibody (A03561-2). RAP1GAP was detected in a paraffin-embedded section of human ovary serous ca
A03561-2 · IFIF analysis of RAP1GAP using anti-RAP1GAP antibody (A03561-2). RAP1GAP was detected in an immunocytochemical section of A549 cells. Enzyme a
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000076864-RAP1GAP/tissue/cerebral+cortex#imid_20754424
HPA · CaudateHuman Protein Atlas IHC, Caudate. Source: https://www.proteinatlas.org/ENSG00000076864-RAP1GAP/tissue/basal+ganglia#imid_20754426
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000076864-RAP1GAP/tissue/colon#imid_20754334
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000076864-RAP1GAP/tissue/liver#imid_20754351
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody CAB003851; green = RAP1GAP, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody CAB003851; green = RAP1GAP, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for RAP1GAP IHC & IF
🧪Run cerebral cortex first: neuronal cells should stain (HPA: High in cerebral cortex neuronal cells). Use adipose tissue adipocytes as the negative comparator (HPA: Not detected in adipocytes); on the cortex slide, assess non-neuronal cells as an internal background reference without assuming they are target-negative.
Positive control tissue: Adrenal gland (Glandular cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show RAP1GAP in A-431, U-251MG, with annotated localisation: Cytosol (approved) (HPA subcellular).
Technical controls: Include a no-primary (secondary-only) control, a concentration-matched rabbit IgG isotype control matched to the primary’s clonality, and knockout material or a validated immunizing-peptide block as a biological specificity control (selected-SKU caption: rabbit primary). For chromogenic staining, quench endogenous peroxidase and inspect the cortex slide for intrinsic pigment before scoring DAB signal (selected-SKU caption: peroxidase/DAB detection).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected-SKU paraffin-section caption does not state the fixative (selected-SKU caption: fixative not stated). The caption uses heat retrieval in EDTA at pH 8.0, but does not establish that retrieval is required or that frozen sections or IF are easier (selected-SKU caption: EDTA heat retrieval). For cortex IF, check an unstained section for tissue autofluorescence before interpreting signal; HPA reports cytosolic ICC-IF localization, while UniProt lists a Golgi membrane association (HPA: Cytosol approved; UniProt P47736 subcellular location).
HPA tissue IHC evidence for RAP1GAP
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 |
|---|---|---|---|---|
| Adrenal gland | Glandular cells | High | Protein (IHC) | HPA → |
| Appendix | Goblet cells | High | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | High | Protein (IHC) | HPA → |
| Caudate | Neuronal cells | High | Protein (IHC) | HPA → |
| Cerebral cortex | Neuronal cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Liver | Cholangiocytes | Not detected | Protein (IHC) | HPA → |
| Lung | Alveolar cells type I | Not detected | Protein (IHC) | HPA → |
| Oral mucosa | Squamous epithelial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced RAP1GAP IHC Tips
Troubleshoot RAP1GAP staining in paraffin sections by checking retrieval, cell identity, subcellular pattern, and assay controls before scoring chromogenic signal.
What retrieval should I start with if RAP1GAP staining is weak?
Start with heat-mediated retrieval in EDTA at pH 8.0 for paraffin sections (datasheet A03561-2). The selected tissue image used this retrieval before staining with 2 μg/ml primary antibody overnight at 4°C, so reproduce that combination before changing one variable at a time (datasheet A03561-2). Check that the section remains attached and that tissue morphology survives heating, because damaged sections can make a weak result difficult to interpret (standard IHC practice). If staining remains weak, compare a second retrieval buffer on adjacent sections while holding antibody concentration and detection constant, and judge both signal and background in the same cell populations (standard IHC practice).
How should I assess whether fixation is suppressing RAP1GAP staining?
Target-specific fixation sensitivity is unknown: the selected RAP1GAP image describes a paraffin section but does not state its fixative (datasheet A03561-2). Record the fixative, fixation duration, processing history, and section age for each specimen before attributing a weak result to the antibody (standard IHC practice). Compare specimens with known processing histories using the same EDTA retrieval at pH 8.0 and 2 μg/ml primary antibody, changing only one processing variable where feasible (datasheet A03561-2; standard IHC practice). Evaluate morphology alongside staining, and include a processed reference section in each run so variation in handling does not masquerade as a biological difference (standard IHC practice).
Should RAP1GAP appear at the Golgi or throughout the cytoplasm?
Score cytoplasmic staining while documenting whether it is diffuse or concentrated near the nucleus: tissue IHC shows cytoplasmic expression, and cellular imaging places RAP1GAP in the cytosol (HPA tissue IHC; HPA subcellular). A Golgi membrane association is also annotated, although RAP1GAP has no transmembrane segment, so morphology alone cannot establish a Golgi-specific signal (UniProt P47736 localisation and topology). Use the same scoring rule across sections and keep nuclear staining separate from cytoplasmic staining in the record (standard IHC practice). If the dominant signal is exclusively nuclear or extracellular, reassess controls, counterstain, and tissue preservation before assigning it to RAP1GAP (HPA tissue IHC; standard IHC practice).
Could isoforms or epitope accessibility explain different staining patterns?
RAP1GAP has 4 annotated isoforms, a GoLoco region at residues 1–17, and a Rap-GAP region at 181–397 (UniProt P47736). The supplied antibody evidence does not locate its epitope, so the staining cannot be assigned to a particular isoform or domain (datasheet A03561-2; UniProt P47736). Several phosphoserines are annotated, including residues 441 and 484, but their effect on this antibody's tissue staining is unestablished (UniProt P47736; datasheet A03561-2). When sections disagree, compare retrieval and processing first, then use an independently validated epitope or isoform-specific assay if that distinction is essential to the experiment (standard IHC practice).
How can I check an IHC pattern with multiplex immunofluorescence?
Use IF as a companion check for cell identity and compartment: pair RAP1GAP with a validated neuronal marker when examining cerebral cortex, where neuronal staining is high (HPA tissue IHC; standard IF practice). Choose spectrally separated fluorophores, preferably placing the weaker signal in a red or far-red channel after checking the specimen's autofluorescence and single-stain controls (standard IF practice). RAP1GAP is reported in the cytosol and at the Golgi membrane without a transmembrane segment, but the supplied evidence does not establish which side contains this antibody's epitope (HPA subcellular; UniProt P47736 topology; datasheet A03561-2). Optimise permeabilisation against an antibody-validated IF control, because the paraffin-section IHC conditions do not establish IF epitope access (standard IF practice; datasheet A03561-2).
What should I change when DAB staining is diffuse or widespread?
First inspect a no-primary control and the tissue edges to separate detection background from a reproducible cellular pattern (standard IHC practice). The selected image used 10% goat serum for blocking, 2 μg/ml primary antibody overnight at 4°C, and a peroxidase-based DAB detection system (datasheet A03561-2). Apply a peroxidase block and assess whether additional washing or lower primary concentration improves contrast without removing staining in expected cells; these are general chromogenic IHC adjustments (standard IHC practice). Compare background with cytoplasmic staining in cerebral cortex neurons or bronchial respiratory epithelial cells, both reported as high, rather than judging the entire section by its darkest area (HPA tissue IHC).
How should I quantify RAP1GAP staining across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring: HPA reports cytoplasmic staining and high signal in cerebral cortex neurons, while some other cell populations are not detected (HPA tissue IHC). For chromogenic sections, record the percentage of positive cells and intensity grades, or calculate an H-score from intensity-weighted percentages on a 0–300 scale (standard IHC practice). Normalise each result to the number of evaluable cells in the predefined compartment, or use positive-cell density per mm² of viable tissue when area is the relevant denominator (standard IHC practice). Keep retrieval, detection, image settings, and scoring thresholds consistent across specimens, and report excluded necrotic or damaged areas (standard IHC practice).
How do I distinguish genuine RAP1GAP staining from an artefact?
A credible result should show cellular cytoplasmic signal in an identified population, consistent with tissue IHC and the approved cytosolic localisation (HPA tissue IHC; HPA subcellular). Cerebral cortex neurons and bronchial respiratory epithelial cells are reported as high, whereas adipocytes and skeletal myocytes are reported as not detected; use these distinctions to assess plausibility, not as absolute controls for every specimen (HPA tissue IHC). Treat staining confined to cut edges, necrotic areas, or the no-primary control as possible processing or detection artefact (standard IHC practice). Investigate predominantly nuclear staining and diffuse DAB deposits with morphology, peroxidase controls, and an independent assay before interpreting them as RAP1GAP biology (HPA subcellular; standard IHC practice).
Boster reagents
Best RAP1GAP / Rap1 GTPase-activating protein 1 IHC Antibodies
A03561-2 has paraffin-section IHC images from human carcinomas and mouse and rat brain, plus an IF image from A549 cells (catalog image captions). M03561 lists IHC and IF/ICC applications (catalog applications).
Real IHC data
Anti-RAP1GAP Antibody ®
Cat # A03561-2
A03561-2 has IHC images from human gastric adenocarcinoma, human ovary serous carcinoma, mouse brain and rat brain paraffin sections, plus an IF image from A549 cells (catalog image captions). M03561 lists IHC and ICC/IF applications and human, mouse and rat reactivity, but provides no IHC or IF image caption (catalog applications, reactivity and image captions).
Which to pick: Choose A03561-2 for paraffin-section IHC: its own captions show staining in human, mouse and rat tissue, and list 2 μg/ml primary antibody with EDTA retrieval at pH 8.0; the fixative is unreported (A03561-2 IHC image captions). Choose A03561-2 for IF/ICC when an image example matters: its own caption shows A549 cells stained at 5 μg/ml (A03561-2 IF image caption). For cross-species work, both SKUs list human, mouse and rat reactivity, but A03561-2 supplies IHC images across all three species; M03561 is a rabbit monoclonal, clone 21R48, with IHC and ICC/IF listed but no image examples supplied (catalog reactivity, clone, applications and image captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P47736 (RPGP1_HUMAN, Rap1 GTPase-activating protein 1).
- Human Protein Atlas. RAP1GAP tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. RAP1GAP subcellular location (ICC-IF): Localized to the cytosol..
- Human Protein Atlas. RAP1GAP antibody validation summary (2 antibodies).
- RAP1GAP is a prognostic biomarker and correlates with immune infiltrates in bladder cancer. Discover oncology 2025 — PMC12098257.
- Downregulation of Rap1GAP through epigenetic silencing and loss of heterozygosity promotes invasion and progression of thyroid tumors. Cancer research 2010 — PMC2822891.
- Low expression of Rap1GAP is associated with epithelial-mesenchymal transition (EMT) and poor prognosis in gastric cancer. Oncotarget 2017 — PMC5352382.
- RAP1GAP inhibits cytoskeletal remodeling and motility in thyroid cancer cells. Endocrine-related cancer 2012 — PMC3531979.
- PubMed PMID:1904317 — UniProt-cited evidence.
- PubMed PMID:9455484 — UniProt-cited evidence.
- PubMed PMID:16710414 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for RAP1GAP (Rap1 GTPase-activating protein 1). Expected localisation Cytoplasmic staining in most tissues (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A03561-2). Validated IHC antibody: A03561-2 at 2-5 μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Glandular cells); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human gastric adenocarcinoma tissue; fixative not specified (datasheet A03561-2); 2. Fixation: Image fixative and duration unreported (datasheet A03561-2); 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: EDTA pH 8.0 (datasheet A03561-2); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A03561-2); 8. Primary antibody: Rabbit anti-RAP1GAP, 2-5 μg/ml (datasheet A03561-2); 9. Primary incubation: Overnight at 4 °C (datasheet A03561-2). 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 A03561-2); Caveat: Antibody staining and RNA expression have medium consistency (HPA tissue IHC); Isoform / epitope: Four isoforms; verify antibody epitope coverage (UniProt); Regulation: Cortex expression exceeds spinal cord (UniProt). Evidence: UniProt P47736, HPA tissue IHC · Enhanced, HPA ICC-IF · cytosol., Datasheet A03561-2, PMC12098257, PMC2822891. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/RAP1GAP/immunohistochemistry-ihc-guide.