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- Table of Contents
This guide uses RAB11A’s cytoplasmic and membranous tissue staining to plan paraffin IHC (HPA tissue IHC). It identifies glandular cells with medium staining and adipocytes reported as unstained for tissue control selection (HPA tissue IHC).
Expected localisation, validated protocols, controls and antibodies — the at-a-glance facts below, then the full design guide.
| Expected localisation | Cytoplasmic and membranous tissue staining (HPA tissue IHC) | |
| Staining pattern | Widespread cytoplasmic and membranous staining (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet M01436-2) | |
| Positive control | Adrenal gland+4 more · see all | |
| Negative control | Adipose tissue+4 more · see all |
| Fixation | Keep fixation consistent across paraffin sections (standard IHC practice; not target-specific) | |
| Caveat | Adipocytes may be unstained despite the broad tissue profile (HPA tissue IHC) | |
| Regulation | Expression regulation is not annotated (UniProt) | |
| Isoform / epitope | 2 isoforms; mature chain spans residues 2–213. Check epitope coverage (UniProt) |
The catalog antibody’s IHC-P protocol is paired with four published RAB11A IHC methods for lung and colon tissues (PMC12376738; PMC6726494; PMC5438609; PMC6605470).
| Sample | Paraffin-embedded human spleen tissue; fixative not specified (datasheet M01436-2) |
| Fixation | Image fixative and duration unreported (datasheet M01436-2); 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 M01436-2); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet M01436-2) |
| Primary antibody | Mouse monoclonal (clone 4H9) anti-RAB11A, 2 μg/ml (datasheet M01436-2) |
| Primary incubation | Overnight at 4 °C (datasheet M01436-2) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet M01436-2) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | RAB11A-positive staining in glandular cells of adrenal gland (HPA tissue IHC: Medium). HPA tissue profile: Ubiquitous cytoplasmic and membranous expression. No signal in the no-primary control. |
RAB11A is associated with recycling endosomes, cytoplasmic vesicles, Golgi membranes and the cell membrane, although it has no transmembrane segment (UniProt P62491 subcellular location; UniProt P62491 topology). In paraffin-section IHC, expect cytoplasmic and membranous staining in selected epithelial cells, including glandular cells in colon and breast (HPA tissue IHC: Medium). HPA describes the tissue pattern as ubiquitous but rates its agreement with RNA expression as medium (HPA tissue IHC: Approved; medium consistency).
| Granular cytoplasmic staining, sometimes concentrated near the nucleus, with some membrane staining in glandular cells. | This fits the reported cytoplasmic and membranous tissue pattern and RAB11A’s vesicle and recycling-endosome locations (HPA tissue IHC; UniProt P62491 subcellular location). In colon or breast glandular cells, compare intensity with HPA’s Medium category; individual cells need not stain identically (HPA tissue IHC; standard IHC interpretation). |
| Strong, uniform nuclear staining with little cytoplasmic or membrane signal. | A predominantly nuclear result is discordant with the supplied RAB11A locations (UniProt P62491 subcellular location; HPA tissue IHC). Review the antibody and detection controls before treating it as a new location; the slide alone cannot identify the cause (standard IHC practice). |
| Prominent staining in a cell population reported as not detected, such as soft-tissue fibroblasts or adipocytes. | This conflicts with those HPA tissue observations and warrants a specificity check (HPA tissue IHC: fibroblasts and adipocytes, Not detected). Cross-reactivity or endogenous detection activity is possible, but biological variation and sampling also limit a single-slide conclusion (standard IHC interpretation). |
| A diffuse wash of color covers cells and surrounding tissue without clear cellular boundaries. | The distribution is hard to reconcile with HPA’s cellular cytoplasmic and membranous pattern (HPA tissue IHC). Excess background can arise from nonspecific antibody binding or the detection system; evaluate a matched negative control before scoring cells (standard IHC practice). |
| No RAB11A signal in a colon or breast glandular-cell section expected to be positive. | HPA reports Medium staining in these cells, so an entirely blank result calls for a run-level check (HPA tissue IHC). Confirm that the section contains the relevant cells and that positive-control tissue and detection reagents worked; HPA’s medium consistency does not guarantee every specimen will stain (HPA tissue IHC; standard IHC practice). |
| Tissue and cell selection | Colon, breast and duodenum glandular cells provide HPA-reported Medium IHC examples; pancreatic exocrine glandular cells are reported as Not detected (HPA tissue IHC). Select and score the named cell population, since a tissue label alone does not specify which cells should stain (standard IHC practice). |
| Antibody evidence | HPA lists IHC as Approved for HPA060686 and CAB013097, while its tissue summary reports medium staining–RNA consistency (HPA antibodies; HPA tissue IHC). Treat agreement with an independent IHC-validated antibody as additional evidence if a disputed pattern must be resolved (standard IHC practice). |
| Membrane association and sequence | RAB11A occupies several membranes despite having no transmembrane segment (UniProt P62491 subcellular location; topology). UniProt lists two isoforms and a short C-terminal propeptide, but the supplied evidence gives no antibody epitope; neither feature predicts this assay’s staining intensity (UniProt P62491 isoforms; processing). |
| IF/ICC Q: What pattern is expected on its separate guide page? | A: HPA supports vesicles and centriolar satellites as the main IF/ICC locations; primary-cilium, basal-body and cytosol assignments are uncertain (HPA subcellular). This localization evidence helps interpret images but does not supply an IF/ICC protocol or establish a paraffin-section staining result (HPA subcellular; HPA tissue IHC). |
| Situation | Likely cause | Next action |
|---|---|---|
| Colon or breast glandular cells and the run’s positive control are both blank. | A shared staining or detection failure is plausible; those glandular cells are HPA-reported Medium examples (HPA tissue IHC; standard IHC practice). | Check section identity, reagent delivery and detection-control performance, then repeat with the catalog antibody’s documented IHC-P conditions (standard IHC practice). |
| The positive control stains, but the chosen study cell population is blank. | Expression can differ by cell population: HPA reports pancreatic exocrine glandular cells as Not detected despite positive glandular examples elsewhere (HPA tissue IHC). | Verify that the scored cells match the HPA comparison and report the negative result for that population; use a separate HPA-positive tissue to assess assay performance (HPA tissue IHC; standard IHC practice). |
| Staining is widespread, including tissue spaces or areas without intact cells. | Nonspecific binding or detection background may obscure the cellular pattern (standard IHC practice). | Compare a matched negative control and inspect blocking, washes and detection settings under the documented IHC-P workflow before scoring (standard IHC practice). |
| Brown signal appears in HPA-reported Not detected cell types. | Specificity is uncertain; cross-reactivity or endogenous chromogen-generating activity may contribute, and an HPA category is not an absolute rule for every specimen (HPA tissue IHC; standard IHC practice). | Review the no-primary control, cell morphology and signal compartment; if the finding matters, compare an independent IHC-validated antibody (standard IHC practice; HPA antibodies). |
| Signal is predominantly nuclear rather than cytoplasmic or membranous. | The compartment conflicts with the supplied tissue and subcellular annotations; these data do not identify which assay step caused it (HPA tissue IHC; UniProt P62491 subcellular location). | Inspect negative controls and repeat with the catalog antibody’s documented IHC-P conditions before assigning a new RAB11A location (standard IHC practice). |
| Granular cytoplasmic staining is uneven across neighboring epithelial cells. | Vesicle-associated localization and differences between cells can produce a nonuniform appearance, but the supplied sources do not define an acceptable cell-by-cell threshold (UniProt P62491 subcellular location; HPA tissue IHC). | Score the named cell population and compartment consistently, recording extent and intensity separately; compare the control section from the same run (standard IHC practice). |
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — 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.
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adrenal gland | Glandular cells | Medium | Protein (IHC) | HPA → |
| Appendix | Glandular cells | Medium | Protein (IHC) | HPA → |
| Breast | Glandular cells | Medium | Protein (IHC) | HPA → |
| Cervix | Squamous epithelial cells | Medium | Protein (IHC) | HPA → |
| Colon | Glandular cells | Medium | Protein (IHC) | HPA → |
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Lymph node | Germinal center cells | Not detected | Protein (IHC) | HPA → |
| Ovary | Ovarian stroma cells | Not detected | Protein (IHC) | HPA → |
| Pancreas | Exocrine glandular cells | Not detected | Protein (IHC) | HPA → |
| Smooth muscle | Smooth muscle cells | Not detected | Protein (IHC) | HPA → |
Troubleshoot RAB11A staining in paraffin sections by checking retrieval, cellular pattern, controls and scoring before interpreting changes in recycling compartments.
Anti-RAB11A antibodies have human paraffin-section IHC and human-cell IF/ICC images (catalog image captions); all three list human, mouse and rat reactivity (catalog reactivity).
M01436-2 has IHC images from human spleen, gastric carcinoma and placenta paraffin sections, plus IF in T-47D cells (M01436-2 image captions); M01436-1 has an IHC-P image from a fixed human brain section (M01436-1 image caption). PB9789 has an IF/ICC image from U2OS cells (PB9789 image caption); all three list human, mouse and rat reactivity (catalog reactivity).
Which to pick: For tissue IHC, choose M01436-2 for paraffin sections with 2 μg/ml primary antibody after EDTA retrieval (M01436-2 IHC captions); its fixative is unreported (M01436-2 IHC captions). M01436-1 is another IHC-P option with a documented paraffin-section human brain section, citrate retrieval and 1:25 primary dilution (M01436-1 image caption); for IF/ICC, choose rabbit PB9789 at 5 μg/ml in the documented U2OS-cell assay (PB9789 image caption). For cross-species planning, all three list human, mouse and rat reactivity (catalog reactivity), while the supplied IHC and IF images show human samples (catalog image captions). The selected M01436-2 tissue-IHC caption documents paraffin sections, but does not specify the fixative (selected-SKU IHC image M01436-2).