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POR / NADPH--cytochrome P450 reductase · IHC design guide
Design Immunohistochemistry for POR
Plan chromogenic POR IHC in paraffin sections using a catalog antibody with documented human tissue staining (datasheet M02166-2). Compare the observed cytoplasmic pattern with HPA tissue IHC, while using POR’s ER membrane location as molecular context (HPA tissue IHC; UniProt).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
POR 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 (HPA tissue IHC); ER membrane (UniProt) | |
| Staining pattern | General cytoplasmic staining, including glandular cells (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 |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent across 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 M02166-2) | |
| Caveat | Medium antibody–RNA concordance complicates interpretation (HPA tissue IHC) | |
| Regulation | Liver-enhanced RNA expression (HPA tissue RNA) | |
| Isoform / epitope | No annotated isoforms; residues 43–677 are cytoplasmic (UniProt) |
Section 1
Recommended POR IHC & IF Protocols
The catalog antibody’s IHC-P protocol is accompanied by one published POR immunohistochemistry protocol in humanized mouse liver (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. |
💡Decision noteStart with the catalog antibody’s heat-mediated EDTA pH 8.0 retrieval (datasheet M02166-2); the published protocol used autoclave retrieval with citrate and EDTA (PMC9437039).
Section 2
What Is the Expected POR Staining Pattern?
POR is an endoplasmic reticulum membrane protein with a short lumenal segment and a predominantly cytoplasmic domain (UniProt P16435 topology). In paraffin tissue sections, expect mainly cytoplasmic staining, including strong staining in adrenal glandular cells and colon glandular cells (HPA tissue IHC). HPA rates the tissue IHC pattern Enhanced, while reporting medium consistency between antibody staining and RNA expression (HPA tissue IHC).
| Cytoplasmic staining in adrenal or colon glandular cells, with a clear counterstained nucleus. | This matches the reported high staining in those cell types and the general cytoplasmic tissue pattern (HPA tissue IHC). The ER membrane assignment supports an intracellular, largely cytoplasmic interpretation (UniProt P16435 topology). Judge intensity against controls processed in the same run (general IHC practice). |
| Predominantly nuclear staining in a paraffin section, with little convincing cytoplasmic signal. | This is discordant with the general tissue IHC pattern and warrants checking controls and staining distribution (HPA tissue IHC; general IHC practice). Do not treat all nuclear signal as disproving POR: HPA also reports supported nucleoplasmic localisation in ICC-IF, a different preparation (HPA subcellular ICC-IF). |
| Strong staining in cardiomyocytes or smooth muscle cells, especially when expected positive cells are weak. | HPA reports POR as not detected in those cell types, so the result raises concern about nonspecific binding or endogenous detection activity (HPA tissue IHC; general IHC practice). It does not alone identify which cause applies; examine reagent controls and morphology (general IHC practice). |
| Broad, uniform colour across cells, stroma, or empty areas that obscures cell boundaries. | This is background rather than a readable cell-specific pattern (general IHC practice). Compare a no-primary control and inspect blocking, washes, and chromogen development before scoring POR; HPA's high and not-detected calls apply to interpretable cellular staining (general IHC practice; HPA tissue IHC). |
| No detectable staining in an adrenal or colon section expected to contain glandular cells. | The absent signal conflicts with HPA's high staining in those cells, but a single negative run cannot establish absent POR (HPA tissue IHC; general IHC practice). Check that the expected cells are present, then assess the positive control and each detection step (general IHC practice). |
💡Expected POR appearanceCall the IHC result positive when intracellular, mainly cytoplasmic colour is clear in adrenal or colon glandular cells at an intensity consistent with HPA's High category; isolated nuclear colour or uniform background is a reason to investigate before calling it POR (HPA tissue IHC; UniProt P16435 topology; general IHC practice).
| Compartment and epitope location | POR has one membrane segment at residues 22–42 and a cytoplasmic region at 43–677 (UniProt P16435 topology). An antibody's epitope is not given here, so topology cannot specify its antigen-retrieval response or predict a unique punctate IHC pattern. |
| Tissue and cell context | HPA reports High staining in adrenal and colon glandular cells, Not detected in cardiomyocytes and smooth muscle cells, and low staining in skeletal myocytes (HPA tissue IHC). Use these as relative interpretation anchors; the HPA summary does not make every cell in a tissue a positive or negative control. |
| Evidence strength | HPA rates the tissue IHC pattern Enhanced and describes medium consistency with RNA expression; HPA010136 and CAB004372 have Enhanced IHC status (HPA tissue IHC; HPA antibodies). These support pattern comparison, while leaving room for sample-to-sample or assay differences. |
| Processing and variants | UniProt lists no isoforms, signal peptide, propeptide, or glycosylation sites, and annotates a 2–677 chain (UniProt P16435). These annotations do not establish a fixation effect or justify attributing an unexpected IHC pattern to a known alternative isoform or shedding event. |
| Situation | Likely cause | Next action |
|---|---|---|
| Expected glandular cells are negative in the same run as other stained tissue. | A failed primary, retrieval, or detection step is possible; the HPA High category does not establish a POR-specific retrieval requirement (HPA tissue IHC; general IHC practice). | Confirm cell identity and section quality, inspect the run's positive control, then check primary dilution, retrieval conditions, and detection reagents against the validated IHC procedure (general IHC practice). |
| Cardiomyocytes or smooth muscle cells show strong colour. | Nonspecific antibody binding or endogenous chromogenic detection activity could mimic a positive signal; HPA reports POR as Not detected in these cells (general IHC practice; HPA tissue IHC). | Compare no-primary and detection-only controls, review morphology, and optimise blocking or endogenous-enzyme quenching where appropriate for the detection system (general IHC practice). |
| Colour covers the section without distinct positive cells. | Excess background can follow insufficient blocking or washing, concentrated reagents, or prolonged chromogen development (general IHC practice). | Use a no-primary control to locate background, then adjust blocking, washes, reagent concentration, or development time one variable at a time (general IHC practice). |
| Signal appears confined to nuclei in paraffin tissue. | The result differs from HPA's general cytoplasmic tissue IHC profile, although supported nucleoplasmic localisation is reported for ICC-IF (HPA tissue IHC; HPA subcellular ICC-IF). | Check cytoplasmic staining in a known positive cell population and review no-primary controls before assigning nuclear IHC signal to POR (HPA tissue IHC; general IHC practice). |
| A negative comparison tissue has faint staining. | Low signal need not equal a failed control: HPA reports low staining in skeletal myocytes, while cardiomyocytes and smooth muscle cells are Not detected (HPA tissue IHC). | Identify the cell type before scoring, compare intensity with the run's positive tissue, and report faint signal separately from strong cell-specific staining (HPA tissue IHC; general IHC practice). |
| Q: Should IF/ICC show the same compartment pattern as paraffin IHC? | HPA tissue IHC reports general cytoplasmic staining; HPA ICC-IF reports mainly vesicles, with additional nucleoplasm and cytosol (HPA tissue IHC; HPA subcellular ICC-IF). | A: Interpret each preparation against its own HPA localisation evidence. Do not use the ICC-IF vesicle call as a required chromogenic IHC pattern (HPA tissue IHC; HPA subcellular ICC-IF). |
Experiment evidence
IHC & IF images showing POR staining
M02166-2 · IHCIHC analysis of POR using anti-POR antibody (M02166-2). POR was detected in a paraffin-embedded section of human esophageal squamous carcino
M02166-2 · IHCIHC analysis of POR using anti-POR antibody (M02166-2). POR was detected in a paraffin-embedded section of human liver cancer tissue. Heat m
PB9736 · IHCIHC analysis of POR using anti-POR antibody (PB9736). POR was detected in paraffin-embedded section of Human Mammary Cancer Tissue. Heat med
PB9736 · IHCIHC analysis of POR using anti-POR antibody (PB9736). POR was detected in paraffin-embedded section of Rat Intestine Tissue. Heat mediated a
PA1952 · IHCAnti-POR antibody, PA1952, IHC(P) IHC(P): Human Placenta Tissue
PA1952 · IHCIHC analysis of POR using anti-POR antibody (PA1952). POR was detected in paraffin-embedded section of rat ovary tissue. Heat mediated antig
PA1952 · IFAnti-POR antibody, PA1952, ICC ICC: HELA Cell
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000127948-POR/tissue/cerebral+cortex#imid_1499940
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000127948-POR/tissue/colon#imid_1499866
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000127948-POR/tissue/liver#imid_1499875
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000127948-POR/tissue/kidney#imid_1499868
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA010136; green = POR, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA010136; green = POR, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for POR IHC & IF
🧪Run adrenal gland first; its glandular cells should stain (HPA: High in adrenal glandular cells). Run heart muscle as the negative tissue (HPA: Not detected in cardiomyocytes); on the adrenal slide, use nonglandular cells as a background comparator, without assuming they are a validated POR-negative population.
Positive control tissue: Adrenal gland (Glandular cells, HPA High)
Negative control tissue: Heart muscle (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show POR in A-431, U-251MG, U2OS, A-549, GAMG, with annotated localisation: Vesicles (supported) (HPA subcellular).
Technical controls: Include no-primary (secondary-only) and concentration-matched mouse IgG isotype controls (selected-SKU IHC caption: mouse anti-POR primary), plus a POR-knockout specimen as a biological negative. Block endogenous peroxidase and check for endogenous biotin background in adrenal tissue when using the caption’s biotin-based, DAB detection method (selected-SKU IHC caption: biotinylated secondary, SABC and DAB).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the exact selected-SKU IHC caption does not state the fixative (selected-SKU IHC caption: fixative unreported). The caption uses heat retrieval in EDTA at pH 8.0, but does not establish that retrieval is required (selected-SKU IHC caption: EDTA retrieval). The supplied evidence does not establish whether frozen sections or IF are easier; for adrenal IHC, evaluate endogenous peroxidase and biotin background with the stated detection method (selected-SKU IHC caption: SABC and DAB).
HPA tissue IHC evidence for POR
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 | 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 → |
Section 3
Advanced POR IHC Tips
Troubleshoot POR staining in paraffin sections by checking retrieval, cell type, compartment, and controls before interpreting signal intensity.
How should I adjust retrieval when POR staining is weak or uneven?
Start with heat-mediated retrieval in EDTA at pH 8.0 (datasheet M02166-2). Use the documented 2 μg/ml primary concentration and overnight incubation at 4°C as a reference when assessing retrieval changes (datasheet M02166-2). If staining remains weak, vary heating duration in matched sections, keeping the antibody concentration, detection, and development time constant (standard IHC practice). Score intact glandular cells alongside the test section, since high POR staining is reported in several glandular tissues (HPA: tissue IHC). Excessive heating can damage morphology or increase diffuse staining; compare cell detail and signal distribution before accepting a stronger result (standard IHC practice).
Could fixation explain weak POR staining in my paraffin sections?
POR-specific sensitivity to fixation is unknown from the supplied evidence; the selected paraffin-section caption does not state a fixative (datasheet M02166-2). Record fixative, fixation duration, section age, and storage conditions for each block before comparing staining intensity (standard IHC practice). Test matched sections from blocks with documented handling, using EDTA retrieval at pH 8.0 and the same detection settings (datasheet M02166-2; standard IHC practice). Check whether weak signal follows a processing batch while tissue architecture and other control stains remain interpretable (standard IHC practice). Do not attribute a fixation effect to POR without a controlled comparison.
Where should POR staining appear, and how should nuclear signal be handled?
Expect predominantly cytoplasmic staining at light-microscope resolution: POR is anchored in the endoplasmic reticulum membrane, with residues 43–677 facing the cytoplasm (UniProt P16435 topology). The tissue IHC profile describes general cytoplasmic expression (HPA: tissue IHC). Cell imaging also reports supported vesicular localisation and additional nucleoplasmic and cytosolic signal (HPA: subcellular). For chromogenic sections, score cytoplasmic staining separately from nuclear staining and compare each pattern with a no-primary control (standard IHC practice). Treat isolated strong nuclear DAB signal cautiously, because the available tissue IHC profile does not establish it as the dominant pattern (HPA: tissue IHC).
How can epitope location change POR staining after retrieval?
The supplied record lists 0 POR isoforms, so an apparent staining difference cannot be assigned to a documented isoform here (UniProt P16435). POR has a membrane-spanning segment at residues 22–42 and a cytoplasmic region at 43–677 (UniProt P16435 topology). It also records acetylation at residue 2 and phosphorylation at residue 63 (UniProt P16435 modified residues). Check the antibody's disclosed immunogen or epitope before relating retrieval response to any of these regions; its epitope is not supplied here. Compare matched sections under the documented EDTA pH 8.0 retrieval conditions before changing staining variables (datasheet M02166-2).
How can IF help resolve an ambiguous POR IHC pattern?
Use IF as a companion check when chromogenic POR staining obscures cell boundaries; the catalog evidence supplied here is a paraffin-section IHC caption (datasheet M02166-2). Multiplex POR with a validated marker for the expected cell type, such as a glandular-cell marker in a glandular tissue (HPA: tissue IHC; standard IF practice). Select fluorophores and image channels after inspecting unstained tissue autofluorescence, with single-stain controls to assess bleed-through (standard IF practice). If the antibody epitope lies within POR residues 43–677, optimise permeabilisation for access to the cytoplasmic side of the endoplasmic reticulum membrane (UniProt P16435 topology; standard IF practice).
What causes widespread brown background in POR IHC sections?
First inspect no-primary and secondary-only controls to distinguish detection-system background from antibody-associated staining (standard IHC practice). In peroxidase-based DAB workflows, block endogenous peroxidase and compare development time across matched sections (standard IHC practice). The documented example used 10% goat serum blocking, a biotinylated secondary antibody, a streptavidin-biotin detection system, and DAB (datasheet M02166-2). With that detection system, consider endogenous biotin as a possible source of background and assess a suitable control (standard IHC practice). Judge residual colour against morphology and the expected predominantly cytoplasmic tissue pattern, rather than treating uniform brown staining as POR signal (HPA: tissue IHC; standard IHC practice).
How should I quantify POR staining across heterogeneous sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the scored cell population before analysis and keep tissue selection, staining batch, and DAB development consistent (standard IHC practice). For glandular regions, report the percentage of positive glandular cells and an H-score combining staining intensity with the percentage at each intensity (standard IHC practice). If spatial distribution matters, report positive cells per mm² of viable, annotated tissue and exclude folds or necrosis (standard IHC practice). Normalise comparisons to the number of eligible cells or viable tissue area, and use a shared reference section to monitor batch variation (standard IHC practice). Report cytoplasmic and nuclear scores separately because general cytoplasmic expression is the tissue IHC pattern (HPA: tissue IHC).
When is a POR-positive DAB pattern biologically credible?
A credible result shows reproducible staining in intact cells, with predominantly cytoplasmic signal consistent with the tissue IHC profile and endoplasmic reticulum membrane localisation (HPA: tissue IHC; UniProt P16435 topology). High staining is reported in adrenal glandular cells and bone-marrow hematopoietic cells, whereas cardiomyocytes and smooth-muscle cells were not detected in the supplied tissue survey (HPA: tissue IHC). Compare the questioned region with those cell-type expectations without assuming every tumour shares normal-tissue expression. Exclude section-edge staining, necrotic deposits, and signal persisting in no-primary or peroxidase controls before calling a positive (standard IHC practice). Interpret borderline results cautiously because antibody staining and RNA show medium consistency in this survey (HPA: Enhanced reliability description).
Boster reagents
Best POR / NADPH--cytochrome P450 reductase IHC Antibodies
Anti-POR IHC images cover paraffin-embedded human carcinoma and placenta sections, plus mouse and rat tissues (catalog image captions: M02166-2, PB9736, PA1952); PA1952 also has a HeLa ICC image (catalog image caption).
Real IHC data
Anti-POR Antibody ® (monoclonal, 7F5)
Cat # M02166-2
Real IHC data
Anti-Cytochrome P450 Reductase/POR Antibody ®
Cat # PB9736
Real IHC data
Anti-Cytochrome P450 Reductase/POR Antibody ®
Cat # PA1952
M02166-2 has IHC images from human esophageal squamous carcinoma, liver cancer, lung cancer and placenta sections; PB9736 has images from human mammary cancer and mouse and rat intestine sections (catalog IHC image captions). PA1952 has IHC images from human placenta and rat ovary sections, plus a HeLa ICC image (catalog image captions).
Which to pick: For human paraffin-section IHC, M02166-2 is a mouse monoclonal with human tissue images (catalog host, clone and IHC image captions). For cross-species IHC, PB9736 lists human, mouse and rat reactivity and has a paraffin-section image for each species (catalog reactivity and IHC image captions). For cell imaging, PA1952 lists ICC and has a HeLa ICC image (catalog applications and image caption); the rendered cards have no IF figure (catalog image captions). The IHC captions document paraffin sections but do not report the fixative (catalog IHC image captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P16435 (NCPR_HUMAN, NADPH--cytochrome P450 reductase).
- Human Protein Atlas. POR tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. POR subcellular location (ICC-IF): Mainly localized to vesicles. In addition localized to the nucleoplasm and cytosol..
- Human Protein Atlas. POR antibody validation summary (3 antibodies).
- Humanized liver TK-NOG mice with functional deletion of hepatic murine cytochrome P450s as a model for studying human drug metabolism. Scientific reports 2022 — PMC9437039.
- Reactivation of CDX2 in Gastric Cancer as Mark for Gene Silencing Memory. Acta histochemica et cytochemica 2015 — PMC4564377.
- Self-Regulated ROS Nanoplatform Enables Sequential Antibacterial Therapy and Inflammation Resolution in Periodontitis. ACS omega 2026 — PMC13557269.
- Integrated follicular fluid multi-omics identifies steroidogenic dysregulation and a candidate SHBG-associated rescue framework in poor ovarian response. Frontiers in endocrinology 2026 — PMC13574599.
- PubMed PMID:1550342 — UniProt-cited evidence.
- PubMed PMID:12853948 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for POR (NADPH--cytochrome P450 reductase). Expected localisation Cytoplasmic staining (HPA tissue IHC); ER membrane (UniProt). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet M02166-2). Validated IHC antibody: M02166-2 at 2 μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Glandular cells); negative Heart muscle. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human esophageal squamous carcinoma tissue; fixative not specified (datasheet M02166-2); 2. Fixation: Image fixative and duration unreported (datasheet M02166-2); 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 M02166-2); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet M02166-2); 8. Primary antibody: Mouse monoclonal (clone 7F5) anti-POR, 2 μg/ml (datasheet M02166-2); 9. Primary incubation: Overnight at 4 °C (datasheet M02166-2). Watch for: Fixation: Keep fixation consistent across 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 M02166-2); Caveat: Medium antibody–RNA concordance complicates interpretation (HPA tissue IHC); Isoform / epitope: No annotated isoforms; residues 43–677 are cytoplasmic (UniProt); Regulation: Liver-enhanced RNA expression (HPA tissue RNA). Evidence: UniProt P16435, HPA tissue IHC · Enhanced, HPA ICC-IF · Mainly localized to vesicles. In addition localized to the…, Datasheet M02166-2, PMC9437039. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/POR/immunohistochemistry-ihc-guide.