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- Table of Contents
Plan POR IHC in paraffin sections using cytoplasmic staining in adrenal gland glandular cells as a positive reference (HPA tissue IHC). This guide covers fixation consistency, an IHC-validated antibody at 2–5 µg/mL (datasheet M02166-2), chromogenic detection, and cell-level scoring.
Expected localisation, validated protocols, controls and antibodies — the at-a-glance facts below, then the full design guide.
| Expected localisation | Cytoplasmic tissue staining (HPA tissue IHC); ER membrane location (UniProt) | |
| Staining pattern | General cytoplasmic staining across cell types (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet M02166-2) | |
| Positive control | Adrenal gland+4 more · see all | |
| Negative control | Heart muscle+1 more · see all |
| Fixation | Keep fixation consistent across sections. (standard IHC practice; not target-specific) | |
| Caveat | Antibody staining and RNA have medium consistency (HPA tissue IHC) | |
| Regulation | Expression regulation not specified (UniProt) | |
| Isoform / epitope | No isoforms annotated; check lumenal vs cytoplasmic epitope (UniProt) |
The catalog antibody protocol uses EDTA pH 8.0 heat retrieval (datasheet: M02166-2). One published POR IHC protocol provides additional fixation, retrieval, detection, and counterstaining details (PMC9437039).
| Sample | Paraffin-embedded human esophageal squamous carcinoma tissue; fixative not specified (datasheet M02166-2) |
| Fixation | Image fixative and duration unreported (datasheet M02166-2); 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 M02166-2); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet M02166-2) |
| Primary antibody | Mouse monoclonal (clone 7F5) anti-POR, 2 μg/ml (datasheet M02166-2) |
| Primary incubation | Overnight at 4 °C (datasheet M02166-2) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet M02166-2) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | POR-positive staining in glandular cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: General cytoplasmic expression. No signal in the no-primary control. |
POR is an endoplasmic reticulum membrane protein with a short lumenal segment and a large cytoplasmic domain (UniProt P16435 topology). In paraffin section IHC, expect a general cytoplasmic staining pattern (HPA tissue IHC). HPA reports high staining in several glandular, respiratory epithelial, neuronal, and hematopoietic cell populations; its tissue profile is rated Enhanced, with medium consistency between staining and RNA expression (HPA tissue IHC).
| Cytoplasmic staining in adrenal glandular cells, with the relevant tissue structures visible. | This fits the reported high glandular cell signal and general cytoplasmic profile (HPA tissue IHC). The large cytoplasmic POR domain makes that compartment plausible (UniProt P16435 topology). Judge intensity against the cells and controls on the same slide; a positive field alone does not establish antibody specificity (general IHC practice). |
| A predominantly nuclear or extracellular deposit, with little cytoplasmic signal. | That is discordant with the tissue IHC profile and ER membrane assignment (HPA tissue IHC; UniProt P16435 subcellular location). Treat it as a possible staining or interpretation artefact and check controls. HPA separately reports nucleoplasmic signal in ICC-IF, so an isolated nuclear IHC result needs validation rather than an automatic biological assignment (HPA subcellular ICC-IF). |
| Strong staining in cardiomyocytes or smooth muscle cells while expected positive cells stain. | HPA reports POR as not detected in those cell types (HPA tissue IHC). Consider cross-reactivity or endogenous chromogen activity, particularly if the deposit does not follow cell boundaries (general IHC practice). Compare an adjacent section without primary antibody before assigning unexpected staining to POR (general IHC practice). |
| A diffuse deposit across cells, stroma, and tissue edges. | This lacks the cell-associated cytoplasmic pattern reported for POR (HPA tissue IHC). Uneven reagent exposure, insufficient blocking or washing, and endogenous detection activity are general IHC possibilities, not documented POR-specific effects (general IHC practice). A primary-omission control can help separate detection background from antibody-dependent staining (general IHC practice). |
| No signal in adrenal glandular cells or another HPA high-staining population. | That conflicts with the reported high staining in the selected positive population (HPA tissue IHC). First confirm that the expected cells are present, then check the IHC-validated antibody, retrieval, detection reagents, and a concurrent positive control (general IHC practice). A failed run cannot establish absent POR expression (general IHC practice). |
| Compartment and epitope orientation | POR has one membrane span at residues 22–42 and a cytoplasmic region at 43–677 (UniProt P16435 topology). The tissue IHC readout is general cytoplasmic staining (HPA tissue IHC). Epitope position for an unspecified antibody is unknown, so topology alone cannot predict its staining strength or retrieval needs. |
| Choice of comparison tissue | HPA reports high staining in adrenal and intestinal glandular cells, respiratory epithelial cells, cortical neuronal cells, and bone marrow hematopoietic cells (HPA tissue IHC). It reports no detection in cardiomyocytes or smooth muscle cells and low staining in skeletal myocytes (HPA tissue IHC). These are useful contrasts, subject to the stated medium staining–RNA consistency (HPA tissue IHC). |
| Interpretation confidence | The tissue profile carries an Enhanced reliability label but only medium consistency between antibody staining and RNA expression (HPA tissue IHC). Two listed antibodies have Enhanced IHC status, while the third has no supplied IHC status (HPA antibodies). These labels support comparison with the reported pattern; they do not validate an unexpected compartment or cell type in a new specimen. |
| Processing and variants | UniProt lists a 2–677 chain, no signal peptide or propeptide, zero annotated glycosylation sites, and zero isoforms (UniProt P16435 processing and isoforms). These annotations give no basis here for a second isoform pattern or a shed, extracellular POR pool. They also do not establish how paraffin processing affects antigen detection. |
| Antigen retrieval | Target-specific fixation effects are not established by the supplied assay evidence. Verify with a matched IHC source before attributing a result to fixation. |
| IF/ICC Q&A: should its pattern match paraffin IHC? | No direct match can be assumed: HPA tissue IHC describes general cytoplasmic expression, whereas HPA ICC-IF reports mainly vesicles, with additional nucleoplasm and cytosol localisation (HPA tissue IHC; HPA subcellular ICC-IF). Those are distinct assay observations. The ICC-IF record does not supply an IHC protocol choice or resolve nuclear-only staining in a paraffin section. |
| Situation | Likely cause | Next action |
|---|---|---|
| Known-positive glandular cells are blank throughout the run. | A run-level staining failure is possible when a population reported as high by HPA is absent (HPA tissue IHC; general IHC practice). | Verify the cell population and concurrent positive control; review retrieval, the IHC-validated antibody, and detection steps using standard paraffin IHC controls (general IHC practice). |
| Only tissue edges or broad acellular regions develop chromogen. | The distribution does not resemble HPA's general cytoplasmic POR profile (HPA tissue IHC); uneven reagent coverage or background is possible (general IHC practice). | Compare a primary-omission section, inspect section coverage, and review blocking and washes before scoring cell-associated signal (general IHC practice). |
| Cardiomyocytes or smooth muscle cells appear strongly positive. | Those cell types are reported as not detected by HPA; antibody cross-reactivity or endogenous detection activity may explain discordant staining (HPA tissue IHC; general IHC practice). | Check a primary-omission control and an HPA high-staining cell population on the same run; investigate the detection system if signal persists without primary antibody (general IHC practice). |
| Staining appears confined to nuclei. | Nuclear-only tissue IHC conflicts with the general cytoplasmic profile (HPA tissue IHC), although nucleoplasm is an additional ICC-IF location (HPA subcellular ICC-IF). | Check the counterstain, compare a primary-omission control, and confirm cytoplasmic staining in a known-positive tissue before interpreting nuclear IHC signal (general IHC practice). |
| A positive tissue stains weakly while other assay controls pass. | The sampled population may differ from the HPA high-staining cell type, or IHC conditions may yield weak signal (HPA tissue IHC; general IHC practice). No POR-specific fixation sensitivity is supplied. | Identify the expected cells first, then compare adjacent controlled sections under documented retrieval and antibody conditions; retain the same scoring threshold across comparisons (general IHC practice). |
| An IF/ICC image looks vesicular but paraffin IHC looks broadly cytoplasmic. | These descriptions come from different HPA assays: vesicles are the main ICC-IF location, while tissue IHC reports general cytoplasmic expression (HPA subcellular ICC-IF; HPA tissue IHC). | Interpret each image against its assay-specific reference and controls; use the separate IF/ICC guide for that assay rather than transferring its localisation pattern into IHC scoring (HPA tissue IHC; HPA subcellular ICC-IF). |
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.
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adrenal gland | Glandular cells | High | Protein (IHC) | HPA → |
| Appendix | Glandular cells | High | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | High | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | High | Protein (IHC) | HPA → |
| Cerebral cortex | Neuronal cells | High | Protein (IHC) | HPA → |
Troubleshoot POR staining in paraffin sections by checking retrieval, compartment, controls and scoring against the reported tissue patterns (datasheet M02166-2; UniProt P16435; HPA tissue IHC).
Anti-POR IHC images cover human esophageal squamous carcinoma, mammary cancer and placenta, plus mouse and rat tissues; an ICC image shows human HeLa cells (catalog image captions).
M02166-2 has human paraffin-section IHC images from esophageal squamous carcinoma, liver cancer, lung cancer and placenta; PB9736 has human mammary cancer and mouse and rat intestine IHC images (catalog image captions). PA1952 has human placenta and rat ovary IHC images plus a human HeLa ICC image (catalog image captions).
Which to pick: For human tissue IHC, M02166-2 is a mouse monoclonal with a paraffin-section image using EDTA retrieval at pH 8.0; the caption does not report the fixative (catalog host/clone; M02166-2 IHC caption). For cross-species tissue IHC, PB9736 lists human, mouse and rat reactivity and has paraffin-section images in all three species using citrate retrieval at pH 6; its captions do not report the fixative (catalog reactivity; PB9736 IHC captions). For IF, M02166 lists IF and human, mouse and rat reactivity but has no IF image; for image-backed ICC, PA1952 has a human HeLa ICC image (catalog applications/reactivity/image alts; PA1952 ICC caption).