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RAB20 / Ras-related protein Rab-20 · IHC design guide
Design Immunohistochemistry for RAB20
Plan RAB20 paraffin-section IHC around cytoplasmic staining reported in several tissues (HPA tissue IHC). Compare positive tissue with controls, while accounting for the low consistency 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
RAB20 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 several tissues (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic staining in glandular cells and hepatocytes (HPA tissue IHC) | |
| Antigen retrieval | Tris-EDTA pH 9.0 HIER, heat-mediated (datasheet A11018) | |
| Positive control | Duodenum+4 more · see all | |
| Negative control | Adipose tissue+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent across sections. (standard IHC practice; not target-specific) | |
| Caveat | Staining has low consistency with RNA expression (HPA tissue IHC) | |
| Regulation | Stronger in pancreatic adenocarcinomas (UniProt) | |
| Isoform / epitope | No isoforms or processing annotated (UniProt) |
Section 1
Recommended RAB20 IHC & IF Protocols
Compare the catalog antibody’s IHC-P protocol (datasheet A11018) with a published RAB20 protocol for paraffin sections of penile squamous cell carcinoma (PMC8909501).
| Sample | Paraffin-embedded human Squamous cell carcinoma of lung tissue; fixative not specified (datasheet A11018) |
| Fixation | Image fixative and duration unreported (datasheet A11018); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat retrieval: Tris-EDTA pH 9.0 (datasheet A11018); 20 min, 95–100 °C (standard) |
| 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-RAB20, 1:100-1:300 (datasheet A11018) |
| 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 | RAB20-positive staining in glandular cells of duodenum (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in several tissues. No signal in the no-primary control. |
💡Decision noteStart with Tris-EDTA pH 9.0 heat retrieval for the catalog antibody (datasheet A11018); the published protocol used citrate pH 6 (PMC8909501).
Section 2
What Is the Expected RAB20 Staining Pattern?
RAB20 is associated with the Golgi and cytoplasmic vesicles, including phagosomes (UniProt Q9NX57 subcellular location); it has no transmembrane segment (UniProt Q9NX57 topology). In paraffin tissue IHC, expect cytoplasmic staining in selected glandular cells, hepatocytes and cardiomyocytes (HPA tissue IHC). Treat tissue intensity as provisional: HPA rates the IHC profile Uncertain because antibody staining has low consistency with RNA expression (HPA tissue IHC).
| Cytoplasmic signal in duodenal or small-intestinal glandular cells, hepatocytes, or cardiomyocytes. | This matches cell types scored High by HPA (HPA tissue IHC). Assess signal within the relevant cells and alongside negative controls; HPA calls the overall tissue IHC profile Uncertain (HPA tissue IHC). |
| Predominantly nuclear staining, or a broad surface outline without convincing cytoplasmic staining. | That distribution conflicts with the reported cytoplasmic tissue profile (HPA tissue IHC) and vesicle/Golgi locations (UniProt Q9NX57 subcellular location). Review controls and detection before assigning it to RAB20 (general IHC practice). |
| Strong signal in bronchial respiratory epithelium or adipocytes while the expected positive cells are weak. | Those cells are scored Not detected (HPA tissue IHC). Consider off-target antibody binding or endogenous detection activity (general IHC practice); the Uncertain HPA rating means staining alone cannot settle specificity (HPA tissue IHC). |
| Color spreads across stroma or extracellular spaces with little separation between cells and background. | Diffuse background obscures the expected cellular cytoplasmic pattern (HPA tissue IHC). Check reagent-only controls, blocking, washes and chromogen development as general IHC troubleshooting; do not score haze as positive cells (general IHC practice). |
| No signal in duodenal glandular cells or hepatocytes in an otherwise interpretable section. | Both are scored High (HPA tissue IHC), so inspect the staining run and its controls before calling the sample negative (general IHC practice). The HPA profile is Uncertain, so these are provisional positive comparators (HPA tissue IHC). |
💡Expected RAB20 appearanceCall a result supportive when cytoplasmic staining is clear in HPA High cell types such as duodenal glandular cells or hepatocytes (HPA tissue IHC); isolated nuclear signal, diffuse haze or strong staining in HPA Not detected cells warrants specificity checks (HPA tissue IHC; general IHC practice).
| Tissue and cell selection | HPA scores duodenal and small-intestinal glandular cells, hepatocytes and cardiomyocytes High; bronchial respiratory epithelium and adipocytes are Not detected (HPA tissue IHC). Compare named cell populations, not whole-section color (general IHC practice). |
| IHC evidence strength | HPA rates the tissue profile Uncertain because staining and RNA expression show low consistency; its listed IHC antibody HPA068647 is also Uncertain (HPA tissue IHC; HPA antibodies). Treat agreement with the pattern as support, not independent proof of specificity. |
| Cellular location and topology | Vesicles, phagosomes and Golgi are reported locations (UniProt Q9NX57 subcellular location); no transmembrane segment is annotated (UniProt Q9NX57 topology). A crisp plasma-membrane-only or nuclear-only result therefore needs review. |
| IF/ICC: what should appear? | The separate IF/ICC guide can assess mainly vesicular signal with additional Golgi signal (HPA subcellular ICC-IF, supported). This cellular localisation evidence can inform interpretation, but does not validate a paraffin IHC staining condition. |
| Processing and fixation evidence | UniProt lists one chain spanning residues 1–234, with no signal peptide or propeptide (UniProt Q9NX57 processing). The supplied sources report no RAB20-specific fixation sensitivity; assess retrieval empirically as general IHC practice. |
| Situation | Likely cause | Next action |
|---|---|---|
| Expected positive cells have no visible signal. | A failed staining step is possible (general IHC practice); duodenal glandular cells and hepatocytes are HPA High comparators, though the profile is Uncertain (HPA tissue IHC). | Check the run control, antibody and detection steps, then optimize retrieval and antibody concentration within the validated IHC workflow (general IHC practice). |
| Nuclei stain strongly while cytoplasm remains weak. | Nuclear dominance conflicts with the cytoplasmic tissue profile (HPA tissue IHC) and vesicle/Golgi locations (UniProt Q9NX57 subcellular location). | Compare with reagent-only and suitable positive controls; repeat with a validated staining setup before interpreting nuclear signal as RAB20 (general IHC practice). |
| Bronchial respiratory epithelial cells stain strongly. | These cells are scored Not detected (HPA tissue IHC); off-target binding or endogenous detection activity is possible (general IHC practice). | Review antibody concentration, blocking and detection controls; compare signal with an HPA High cell population on the same run (HPA tissue IHC; general IHC practice). |
| Diffuse chromogen obscures cell boundaries. | Background may reflect nonspecific reagent binding, incomplete washes or excessive development (general IHC practice). | Inspect reagent-only controls and adjust blocking, washes or development within the assay workflow; score only distinguishable cellular staining (general IHC practice). |
| Liver or another highly colored section shows widespread signal. | Endogenous detection activity or tissue pigment can mimic chromogenic staining (general IHC practice); hepatocytes are HPA High, so color alone is insufficient (HPA tissue IHC). | Use appropriate detection and negative controls, examine the expected cellular distribution, and distinguish pigment from reaction product (general IHC practice). |
| A plausible cytoplasmic pattern differs between sections or antibodies. | HPA reports low consistency between antibody staining and RNA expression, and rates tissue IHC Uncertain (HPA tissue IHC). | Compare matched cell types and controls across runs; document the disagreement and seek independent specificity evidence before making a firm biological claim (general IHC practice). |
Experiment evidence
IHC & IF images showing RAB20 staining
A11018 · IHCImmunohistochemical analysis of paraffin-embedded human Squamous cell carcinoma of lung. 1, Antibody was diluted at 1:200 (4° overnight). 2,
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000139832-RAB20/tissue/cerebral+cortex#imid_18832562
HPA · Adrenal glandHuman Protein Atlas IHC, Adrenal gland. Source: https://www.proteinatlas.org/ENSG00000139832-RAB20/tissue/adrenal+gland#imid_18832543
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000139832-RAB20/tissue/colon#imid_18742086
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000139832-RAB20/tissue/liver#imid_18742087
HPA · ICC-IF CACO-2Human Protein Atlas immunofluorescence, CACO-2, antibody HPA002882; green = RAB20, blue = nucleus, red = microtubules.
HPA · ICC-IF CACO-2Human Protein Atlas immunofluorescence, CACO-2, antibody HPA002882; green = RAB20, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for RAB20 IHC & IF
🧪Run duodenum first; glandular cells should stain (HPA: High in duodenal glandular cells). Use adipose tissue as the negative comparator (HPA: Not detected in adipocytes); on the duodenum slide, assess adjacent nonglandular cells for background without assuming they are RAB20-negative (HPA: only glandular cells specified).
Positive control tissue: Duodenum (Glandular cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show RAB20 in CACO-2, SK-MEL-30, U2OS, A-431, with annotated localisation: Vesicles (supported) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control and an isotype control matched to the primary antibody’s host species and immunoglobulin class (standard IHC practice); use a RAB20 knockout specimen as a biological specificity control if available (standard IHC practice). For chromogenic staining, block endogenous peroxidase and check background from any biotin-based detection system (standard IHC practice).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected A11018 tissue-IHC caption does not state the fixative (selected-SKU caption: fixative unreported). The paraffin-section example uses Tris-EDTA at pH 9.0 for retrieval and primary antibody at 1:200 overnight at 4°C, but it does not establish whether retrieval is essential (selected-SKU caption). The supplied evidence does not establish that frozen sections or IF are easier; vesicular and Golgi localization makes spatial interpretation relevant for IF (HPA subcellular: vesicles and Golgi apparatus).
HPA tissue IHC evidence for RAB20
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Uncertain — 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 |
|---|---|---|---|---|
| Duodenum | Glandular cells | High | Protein (IHC) | HPA → |
| Heart muscle | Cardiomyocytes | High | Protein (IHC) | HPA → |
| Liver | Hepatocytes | High | Protein (IHC) | HPA → |
| Small intestine | Glandular cells | High | Protein (IHC) | HPA → |
| Adrenal 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 → |
| 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 → |
| Cerebellum | Cells in granular layer | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced RAB20 IHC Tips
Use the documented paraffin section conditions as a starting point, then verify RAB20 staining by compartment, cell type, and controls.
Which retrieval conditions should I start with for RAB20 paraffin sections?
Start with heat-mediated Tris-EDTA retrieval at pH 9.0 for paraffin sections stained with the catalog antibody (datasheet A11018). Pair it with the documented primary antibody dilution of 1:200 and incubation at 4°C overnight before changing several variables at once (datasheet A11018). If staining is weak, compare retrieval heating times on adjacent sections while keeping the antibody and detection conditions constant (standard IHC practice). Assess whether any increase reveals cytoplasmic vesicular staining without simultaneously increasing diffuse background or damaging tissue morphology; vesicles and the Golgi are supported RAB20 locations (HPA subcellular; standard IHC practice).
How should I troubleshoot fixation-related loss of RAB20 staining?
The selected tissue image documents paraffin-embedded lung squamous cell carcinoma but does not state its fixative, so target-specific fixation sensitivity is unknown (datasheet A11018). Record each specimen’s fixative, fixation duration, and processing history, then compare similarly processed sections before attributing a staining difference to RAB20 abundance (standard IHC practice). If staining varies across a block, examine morphology and compare interior with peripheral tissue while holding Tris-EDTA pH 9.0 retrieval constant (datasheet A11018; standard IHC practice). Use a known staining control processed alongside the samples, and report any fixation differences as a limitation of the comparison (standard IHC practice).
What staining pattern is consistent with RAB20 in chromogenic IHC?
Look for cytoplasmic staining with a vesicular distribution; RAB20 is mainly localized to vesicles and additionally to the Golgi apparatus (HPA subcellular). RAB20 is also associated with phagosomes and their membranes, and the record reports no transmembrane segment, so a sharp, continuous plasma membrane outline needs independent validation (UniProt Q9NX57). In paraffin sections, judge puncta against cell boundaries and morphology at the same magnification used for controls, because chromogenic signal can obscure small structures (standard IHC practice). Do not require every positive cell to show resolvable puncta, but investigate dominant nuclear or extracellular staining before scoring it as RAB20 (HPA subcellular; standard IHC practice).
How can I assess whether the antibody detects the intended RAB20 epitope?
The supplied record lists a single 1–234 protein chain, no annotated isoforms, no transmembrane segment, and no annotated glycosylation sites or modified residues (UniProt Q9NX57). Those annotations cannot establish which epitope the catalog antibody recognizes or whether processing exposes it in paraffin sections (UniProt Q9NX57; standard IHC practice). Check the antibody’s stated immunogen or epitope information, if available, and compare staining after the documented Tris-EDTA pH 9.0 retrieval with appropriate controls (datasheet A11018; standard IHC practice). If specificity remains uncertain, seek an independent antibody recognizing a different epitope and require concordant cellular and subcellular patterns before interpreting differences biologically (standard IHC practice).
How should I use IF to investigate an ambiguous RAB20 IHC pattern?
Use IF as a separate validation experiment rather than transferring the paraffin section antibody conditions directly; the selected caption reports tissue IHC conditions only (datasheet A11018). Multiplex RAB20 with a marker for the expected cell population, choosing that marker from the specimen’s verified cell identity, and inspect whether signal falls within those cells (standard IF practice). Choose fluorophores and image channels after checking tissue autofluorescence with an unstained control, particularly when weak puncta are being assessed (standard IF practice). Because RAB20 is associated with intracellular vesicles and phagosome membranes, optimize permeabilisation for access to the antibody’s mapped epitope; its epitope orientation is not supplied here (UniProt Q9NX57; standard IF practice).
How can I distinguish nonspecific brown staining from RAB20 signal?
Run a no-primary control through the same chromogenic detection workflow to identify signal from the detection reagents or endogenous tissue activity (standard IHC practice). Include a peroxidase block when using a peroxidase and DAB workflow, and inspect pigment, folds, and section edges before calling brown deposits positive (standard IHC practice). The documented secondary antibody condition is 1:200 for 45 minutes at room temperature; keep it fixed while adjusting primary antibody exposure or blocking if background persists (datasheet A11018; standard IHC practice). Prefer intracellular staining consistent with RAB20’s vesicular and Golgi locations over diffuse staining that also appears in the no-primary control (HPA subcellular; standard IHC practice).
What is a defensible way to quantify RAB20 IHC across samples? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and analysis regions before scoring, then use percent positive cells and an intensity-based H-score within that population (standard IHC practice). Record both the numerator and the number of evaluable target cells, and exclude necrosis, folds, and damaged edges using the same rules for every section (standard IHC practice). For a spatial analysis, report positive-cell density per mm² of viable tissue and normalize comparisons to the same cell compartment or tissue area definition (standard IHC practice). Keep retrieval, imaging, and scoring thresholds consistent; HPA describes its tissue IHC reliability as uncertain because antibody staining and RNA expression have low consistency (datasheet A11018; HPA tissue IHC).
When should I distrust an apparent RAB20-positive IHC result?
Question signal dominated by nuclei, extracellular material, necrosis, or section edges, since the supported locations are intracellular vesicles and the Golgi apparatus (HPA subcellular; standard IHC practice). Check whether staining follows plausible cell boundaries and persists beyond areas with pigment or endogenous enzyme signal in the no-primary control (standard IHC practice). HPA reports cytoplasmic expression in several tissues but rates its tissue IHC evidence uncertain because staining has low consistency with RNA expression, so a single stained section is insufficient for a strong biological conclusion (HPA tissue IHC). Compare matched controls and, when feasible, an independent specificity method before interpreting between-sample differences as altered RAB20 expression (standard IHC practice).
Boster reagents
Best RAB20 / Ras-related protein Rab-20 IHC Antibodies
A11018 has a real IHC image from a paraffin-embedded human lung squamous cell carcinoma section (image caption); IF is listed, with Human and Mouse reactivity (catalog).
Real IHC data
Anti-Rab 20 Antibody
Cat # A11018
A11018 will render with its paraffin-section IHC image of human lung squamous cell carcinoma (image caption). The catalog lists IHC and IF applications and Human and Mouse reactivity for A11018 (catalog); no IF image is supplied (catalog).
Which to pick: Choose A11018 for paraffin-section IHC: its image caption reports primary antibody at 1:200 overnight at 4°C and Tris-EDTA retrieval at pH 9.0 (image caption). A11018 is also the listed IF/ICC option because IF appears in its application list, although no IF image is supplied (catalog). For Human or Mouse samples, A11018 is the only listed option (catalog reactivity); its host is Rabbit and clonality is unreported (catalog), while the IHC caption does not report a fixative (image caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q9NX57 (RAB20_HUMAN, Ras-related protein Rab-20).
- Human Protein Atlas. RAB20 tissue IHC expression (reliability: Uncertain).
- Human Protein Atlas. RAB20 subcellular location (ICC-IF): Mainly localized to vesicles. In addition localized to the Golgi apparatus..
- Human Protein Atlas. RAB20 antibody validation summary (3 antibodies).
- RAB20 Promotes Proliferation via G2/M Phase through the Chk1/cdc25c/cdc2-cyclinB1 Pathway in Penile Squamous Cell Carcinoma. Cancers 2022 — PMC8909501.
- PubMed PMID:16613320 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
- PubMed PMID:15057823 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for RAB20 (Ras-related protein Rab-20). Expected localisation Cytoplasmic staining in several tissues (HPA tissue IHC). Antigen retrieval: Tris-EDTA pH 9.0 HIER, heat-mediated (datasheet A11018). Validated IHC antibody: A11018 at 1:100-1:300 (the product's own IHC validation image is shown). Controls (HPA): positive Duodenum (Glandular cells); negative Adipose tissue. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Paraffin-embedded human Squamous cell carcinoma of lung tissue; fixative not specified (datasheet A11018); 2. Fixation: Image fixative and duration unreported (datasheet A11018); 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: Tris-EDTA pH 9.0 (datasheet A11018); 20 min, 95–100 °C (standard); 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-RAB20, 1:100-1:300 (datasheet A11018); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across sections. (standard IHC practice; not target-specific); Caveat: Staining has low consistency with RNA expression (HPA tissue IHC); Isoform / epitope: No isoforms or processing annotated (UniProt); Regulation: Stronger in pancreatic adenocarcinomas (UniProt). Evidence: UniProt Q9NX57, HPA tissue IHC · Uncertain, HPA ICC-IF · Mainly localized to vesicles. In addition localized to the Golgi…, Datasheet A11018, PMC8909501. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/RAB20/immunohistochemistry-ihc-guide.