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ENTPD5 / Nucleoside diphosphate phosphatase ENTPD5 · IHC design guide
Design Immunohistochemistry for ENTPD5
Plan ENTPD5 IHC in paraffin sections using hepatocytes and kidney proximal tubules as positive controls (HPA tissue IHC). This guide covers fixation, antibody dilution and chromogenic scoring with the IHC-validated antibody (datasheet A06908-2).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
ENTPD5 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 tissue staining (HPA tissue IHC) | |
| Staining pattern | Strong cytoplasmic staining in hepatocytes and proximal tubules (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A06908-2) | |
| Positive control | Appendix+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 | A secreted variant may shift tissue staining (HPA tissue IHC) | |
| Regulation | No expression regulator annotated (UniProt) | |
| Isoform / epitope | No isoforms annotated; signal peptide 1–24 is cleaved (UniProt) |
Section 1
Recommended ENTPD5 IHC & IF Protocols
The catalog antibody’s IHC-P protocol (datasheet A06908-2) is accompanied by two published ENTPD5 paraffin-section protocols (PMC3743121; PMC4368616).
| Sample | Paraffin-embedded human lung cancer tissue; fixative not specified (datasheet A06908-2) |
| Fixation | Image fixative and duration unreported (datasheet A06908-2); 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 A06908-2); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A06908-2) |
| Primary antibody | Rabbit anti-ENTPD5, 2-5 μg/ml (datasheet A06908-2) |
| Primary incubation | Overnight at 4 °C (datasheet A06908-2) |
| Detection | HRP-conjugated secondary, DAB chromogen (datasheet A06908-2) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | ENTPD5-positive staining in endocrine cells of appendix (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in most tissues, highly expressed in liver, kidney, prostate and gastrointestinal mucosa. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA pH 8.0 retrieval for the catalog antibody (datasheet A06908-2); neither published excerpt specifies retrieval (PMC3743121; PMC4368616).
Section 2
What Is the Expected ENTPD5 Staining Pattern?
Expect mainly cytoplasmic ENTPD5 staining in hepatocytes, kidney proximal tubules, and selected gastrointestinal cells (HPA: tissue IHC, Enhanced reliability). UniProt places ENTPD5 in the endoplasmic reticulum and also annotates it as secreted; it has no transmembrane segment (UniProt O75356: subcellular location and topology). Interpret cell identity and staining distribution together, since a secreted variant can complicate tissue-level agreement between RNA and protein (HPA: reliability description).
| Strong cytoplasmic staining in hepatocytes and kidney proximal tubules, with recognizable tissue structure. | This matches two high-staining cell populations (HPA: High in hepatocytes and proximal tubules). Compare intensity within the intended cell population rather than treating all cells in a positive tissue as equivalent (general IHC practice). |
| Predominantly nuclear staining, or a sharp membrane outline with little cytoplasmic signal. | The distribution conflicts with HPA’s predominantly cytoplasmic tissue profile and UniProt’s ER location and lack of a transmembrane segment (HPA: tissue IHC; UniProt O75356: location and topology). Check the counterstain and antibody specificity before scoring it as ENTPD5 (general IHC practice). |
| Strong reaction product in cells expected to be unstained, while expected positive cells are weak. | For example, strong staining of cardiomyocytes or adipocytes conflicts with their HPA not-detected calls (HPA: heart muscle cardiomyocytes; adipose adipocytes). Cross-reactivity or endogenous detection activity is possible; investigate with appropriate staining controls (general IHC practice). |
| Haze or precipitate extends across cells and surrounding tissue without a coherent cellular pattern. | Diffuse background cannot establish ENTPD5 expression by itself (general IHC practice). Because UniProt also annotates ENTPD5 as secreted, extracellular signal needs careful interpretation rather than automatic dismissal; compare it with cellular staining and controls (UniProt O75356: secreted; general IHC practice). |
| Little or no staining in a section containing hepatocytes or kidney proximal tubules. | That is discordant with the high HPA staining reported for those cells (HPA: liver hepatocytes; kidney proximal tubules). First assess tissue preservation, detection performance, and whether the expected cells are present on the examined section (general IHC practice). |
💡Expected ENTPD5 appearanceCall positive when hepatocytes or kidney proximal tubules show clear, high cytoplasmic staining (HPA: High in both); dominant nuclear staining or strong signal in HPA not-detected cells warrants a specificity check (HPA: tissue IHC; general IHC practice).
| Cell population and tissue choice | HPA reports high staining in liver hepatocytes, kidney proximal tubules, and endocrine cells of appendix, colon, and duodenum (HPA: tissue IHC). It reports low staining in lung macrophages and no detection in cardiomyocytes, so these populations have different interpretive value (HPA: tissue IHC). |
| Location and secreted variant | An ER-associated protein can yield cytoplasmic tissue staining, while the secreted annotation makes extracellular signal possible to consider (UniProt O75356: location; HPA: cytoplasmic profile). HPA cautions that RNA and protein locations may differ for a secreted variant (HPA: reliability description). |
| Processing and molecular features | UniProt annotates a signal peptide at residues 1–24, a mature chain at 25–428, and glycosylation sites at 232 and 368 (UniProt O75356: processing and glycosylation). The supplied sources do not locate the antibody epitope, so these features cannot predict its staining sensitivity. |
| Antibody evidence | The HPA tissue profile has Enhanced reliability, and HPA lists antibody HPA002927 as IHC Enhanced (HPA: tissue IHC and antibody validation). That supports using its observed pattern as a comparison, while an individual slide still needs appropriate controls (general IHC practice). |
| IF/ICC Q&A: Is a punctate intracellular IF pattern established? | No defined main ICC-IF location or cell-line images are provided (HPA: subcellular record). UniProt annotates ER and secreted locations, but those annotations alone do not validate a particular IF pattern (UniProt O75356: location; HPA: subcellular record). |
| Situation | Likely cause | Next action |
|---|---|---|
| Known-positive liver or kidney appears negative. | The result conflicts with high staining in hepatocytes and proximal tubules (HPA: tissue IHC); the precise technical cause cannot be identified from this record. | Confirm that those cells are present, inspect section quality, and check antibody, detection, and control performance (general IHC practice). Optimize retrieval only as a general IHC variable; no ENTPD5-specific retrieval or fixation sensitivity is supplied. |
| Signal is mostly nuclear. | A nuclear-dominant result disagrees with the cytoplasmic HPA profile and ER annotation (HPA: tissue IHC; UniProt O75356: location). | Review the unstained or counterstained appearance, compare a no-primary control, and reassess whether the signal follows the expected positive cells (general IHC practice). |
| Unexpected cells stain more strongly than expected positive cells. | This may reflect antibody cross-reactivity or endogenous detection activity (general IHC practice). HPA reports cardiomyocytes, adipocytes, and fibroblasts as not detected (HPA: tissue IHC). | Verify cell identity and include suitable no-primary and detection controls; address endogenous enzyme activity if relevant to the chromogenic system (general IHC practice). |
| Weak staining occurs throughout nearly every compartment. | Widespread low-level reaction product may reflect background rather than a cell-specific pattern (general IHC practice). HPA describes cytoplasmic expression in most tissues, so weak staining alone is not decisive (HPA: tissue IHC). | Compare with no-primary background and a high-staining reference cell population; review blocking, antibody concentration, washing, and detection conditions (general IHC practice). |
| Extracellular material stains, but cellular staining is uncertain. | UniProt annotates ENTPD5 as secreted as well as ER-localized (UniProt O75356: location). HPA warns that a secreted variant can complicate RNA–protein tissue correlation (HPA: reliability description). | Record extracellular and cellular signal separately, compare matched controls, and avoid assigning the extracellular deposit to a producing cell without supporting evidence (general IHC practice). |
| A presumed negative tissue shows scattered positive cells. | A whole-tissue label can obscure cell-specific differences: HPA reports low lung macrophage staining despite low or absent calls in other listed cell populations (HPA: tissue IHC). | Identify the stained cell type before calling the tissue positive or nonspecific; compare that cell type with the corresponding HPA entry and assess controls (HPA: tissue IHC; general IHC practice). |
Experiment evidence
IHC & IF images showing ENTPD5 staining
A06908-2 · IHCIHC analysis of ENTPD5 using anti-ENTPD5 antibody (A06908-2). ENTPD5 was detected in a paraffin-embedded section of human lung cancer tissue
A06908-2 · IHCIHC analysis of ENTPD5 using anti-ENTPD5 antibody (A06908-2). ENTPD5 was detected in a paraffin-embedded section of human lung cancer tissue
A06908-2 · IFIF analysis of ENTPD5 using anti-ENTPD5 antibody (A06908-2). ENTPD5 was detected in a paraffin-embedded section of human lung cancer tissue.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000187097-ENTPD5/tissue/cerebral+cortex#imid_941176
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000187097-ENTPD5/tissue/colon#imid_941086
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000187097-ENTPD5/tissue/liver#imid_941103
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000187097-ENTPD5/tissue/kidney#imid_941088
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for ENTPD5 IHC & IF
🧪Run colon first and look for staining in endocrine cells (High; HPA: colon endocrine cells). Use heart muscle as the negative tissue: cardiomyocytes are Not detected (HPA: heart muscle); assess neighboring non-endocrine cells on the colon slide as internal background comparators without assuming they are ENTPD5-negative (standard IHC practice).
Positive control tissue: Appendix (Endocrine cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA carries no ICC-IF cell line for ENTPD5; derive a cell-line control from the positive tissue's cell type (Endocrine cells) and confirm it by RNA or western blot first.
Technical controls: Include no-primary (secondary-only) and concentration-matched rabbit isotype controls, plus ENTPD5 knockout tissue or a validated peptide-block control if available (selected-SKU caption: rabbit primary antibody; standard IHC practice). Quench endogenous peroxidase and inspect the colon section for nonspecific DAB staining (selected-SKU caption: HRP/DAB detection; standard IHC practice).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected-SKU tissue-IHC caption does not state a fixative (selected-SKU caption: fixative unreported). The reported paraffin-section example used heat retrieval in EDTA at pH 8.0, but retrieval dependency has not been established (selected-SKU caption: EDTA retrieval). Frozen-section or IF ease cannot be ranked from the supplied evidence; HPA lists no ICC-IF cell-line images, and colon background should be checked with the proposed controls (HPA: no ICC-IF cell-line images; standard IHC practice).
HPA tissue IHC evidence for ENTPD5
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Enhanced — High consistency between antibody staining and RNA expression data. At least one protein variant secreted, tissue location of RNA and protein might differ and correlation is complex.). 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 |
|---|---|---|---|---|
| Appendix | Endocrine cells | High | Protein (IHC) | HPA → |
| Breast | Glandular cells | High | Protein (IHC) | HPA → |
| Bronchus | Ciliated cells (cell body) | High | Protein (IHC) | HPA → |
| Colon | Endocrine cells | High | Protein (IHC) | HPA → |
| Duodenum | Endocrine cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Heart muscle | Cardiomyocytes | Not detected | Protein (IHC) | HPA → |
| Ovary | Ovarian stroma cells | Not detected | Protein (IHC) | HPA → |
| Skeletal muscle | Myocytes | Not detected | Protein (IHC) | HPA → |
| Smooth muscle | Smooth muscle cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced ENTPD5 IHC Tips
Use the catalog antibody’s paraffin-section IHC conditions as the starting point, then check staining against ENTPD5’s reported cell and compartment patterns.
Which retrieval condition should I start with for ENTPD5 paraffin-section IHC?
Start with heat-mediated retrieval in EDTA at pH 8.0 for paraffin sections (datasheet A06908-2). The selected tissue-IHC caption reports this condition, followed by 2 μg/ml primary antibody overnight at 4°C; it does not state the fixative (datasheet A06908-2). If staining is weak, vary heating duration on adjacent sections while keeping buffer and detection conditions constant (standard IHC practice). Compare each run with a positive hepatocyte or kidney proximal-tubule control and a no-primary control (HPA: High in hepatocytes and kidney proximal tubules; standard IHC practice). Excessive tissue damage or diffuse staining calls for gentler retrieval before increasing antibody concentration (standard IHC practice).
How should I troubleshoot fixation-related loss of ENTPD5 staining?
The selected paraffin-section caption does not report its fixative, so target-specific fixation sensitivity is unknown (datasheet A06908-2). Record fixative, time before fixation, fixation duration and section age for each specimen before comparing staining intensity (standard IHC practice). If a known positive control stains inconsistently, process control and test tissue together, then compare adjacent sections using the stated EDTA pH 8.0 retrieval condition (datasheet A06908-2; standard IHC practice). Assess morphology and background alongside signal, because stronger retrieval can expose epitopes while damaging sections or increasing nonspecific staining (standard IHC practice). Neither reported tissue expression nor ENTPD5 topology establishes a preferred fixative for this antibody (HPA: tissue IHC; UniProt O75356 topology).
What ENTPD5 staining pattern is plausible in a paraffin section?
Expect predominantly cytoplasmic staining in many tissues, while allowing for context-dependent extracellular signal because ENTPD5 is reported in the endoplasmic reticulum and as secreted (HPA: tissue IHC profile; UniProt O75356 subcellular location). It has no annotated transmembrane segment, so a crisp cell-surface rim alone deserves scrutiny (UniProt O75356 topology). Hepatocytes and kidney proximal tubules provide reported strongly staining cell populations for comparison (HPA: High in hepatocytes and kidney proximal tubules). Examine whether signal follows intact cell bodies rather than folds, cut edges or necrotic material (standard IHC practice). Interpret extracellular deposits cautiously with a no-primary control and adjacent tissue morphology, since location alone cannot establish specificity for a secreted protein (UniProt O75356 subcellular location; standard IHC practice).
How do processing and glycosylation affect ENTPD5 epitope interpretation?
ENTPD5 has no annotated isoforms in the supplied record, but its signal peptide spans residues 1–24 and the mature chain spans 25–428 (UniProt O75356 processing and isoforms). Glycosylation is annotated at residues 232 and 368, which makes epitope position relevant when evaluating access after processing (UniProt O75356 glycosylation; standard IHC practice). The supplied caption does not map this antibody’s epitope, so do not assume it recognizes the signal peptide, mature chain or a glycosylation-sensitive site (datasheet A06908-2). Request an epitope map if staining varies across preparations, and compare matched sections under the stated EDTA pH 8.0 retrieval condition (datasheet A06908-2; standard IHC practice). A staining difference by itself does not establish an ENTPD5 isoform (UniProt O75356 isoforms; standard IHC practice).
How should I assess ENTPD5 by multiplex IF after seeing chromogenic IHC staining?
Treat IF as a separate assay: the selected antibody evidence describes chromogenic staining in a paraffin section, not an IF result (datasheet A06908-2). Multiplex ENTPD5 with a marker identifying the expected cell population, such as hepatocytes or kidney proximal tubules, and inspect each channel separately (HPA: High in hepatocytes and kidney proximal tubules; standard IF practice). Choose spectrally separated fluorophores after examining unstained tissue autofluorescence, which can obscure weak signal (standard IF practice). Because ENTPD5 can reside in the ER lumen and the antibody’s epitope is unmapped here, compare permeabilised and minimally permeabilised preparations before interpreting intracellular absence (UniProt O75356 subcellular location; datasheet A06908-2; standard IF practice). Include single-label and no-primary controls when judging colocalisation (standard IF practice).
How can I separate ENTPD5 signal from chromogenic background?
First inspect a no-primary section for staining from the detection system, then compare it with the primary-stained section using identical development conditions (standard IHC practice). The selected method uses 10% goat serum blocking, 2 μg/ml primary antibody and an HRP-linked anti-rabbit detection workflow with DAB (datasheet A06908-2). Apply a peroxidase block and check the no-primary control when tissue-associated peroxidase activity could generate DAB signal (standard IHC practice). If background persists, shorten DAB development or titrate the primary on adjacent sections while preserving a positive control (standard IHC practice). Judge granular deposits against tissue morphology; staining at folds and damaged edges requires particular caution (standard IHC practice).
What should ENTPD5 IHC scores measure across specimens? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and cytoplasmic compartment before scoring, because the reported tissue pattern is largely cytoplasmic and differs by cell type (HPA: tissue IHC profile). For an H-score, record the percentage of eligible cells at intensity 0–3 and sum percentage multiplied by intensity, yielding 0–300 (standard IHC scoring practice). Also report the percentage of positive eligible cells so intensity shifts remain distinguishable from changes in cell abundance (standard IHC scoring practice). Normalise to the number of assessable target cells or their measured tissue area, excluding necrosis and folds by a prespecified rule (standard IHC scoring practice). Keep retrieval, development and image settings consistent, and score a shared control section across batches (standard IHC practice).
When is an apparent ENTPD5-positive IHC result convincing?
A convincing result combines plausible cytoplasmic staining with expected cell populations, such as hepatocytes or kidney proximal tubules, and a clean no-primary control (HPA: cytoplasmic profile and High in hepatocytes and kidney proximal tubules; standard IHC practice). Interpret staining outside cells cautiously because ENTPD5 is also reported as secreted (UniProt O75356 subcellular location). A purely nuclear pattern, strong cut-edge signal or staining concentrated in necrotic material needs further validation against morphology and controls (UniProt O75356 subcellular location; standard IHC practice). Check for endogenous peroxidase signal when DAB appears without primary antibody (standard IHC practice). HPA reports enhanced agreement between antibody staining and RNA expression, while noting that secretion complicates their spatial comparison (HPA: Enhanced reliability description).
Boster reagents
Best ENTPD5 / Nucleoside diphosphate phosphatase ENTPD5 IHC Antibodies
Anti-ENTPD5 IHC images cover paraffin sections of human lung cancer, rat kidney, and mouse adrenal tissue; an IF image covers human lung cancer sections (A06908-2 image captions).
Real IHC data
Anti-ENTPD5 Antibody ®
Cat # A06908-2
A06908-2 has IHC images from human lung cancer, rat kidney, and mouse adrenal paraffin sections, plus an IF image from human lung cancer paraffin sections (A06908-2 image captions). M06908-2 lists IHC and human, mouse, and rat reactivity, but the payload provides no IHC or IF image for it (M06908-2 catalog applications, reactivity, and image alts).
Which to pick: For tissue IHC, choose A06908-2: its own paraffin-section captions document EDTA retrieval at pH 8.0 and 2 μg/ml primary antibody; the fixative is unreported (A06908-2 IHC image captions). For IF, choose A06908-2 based on its human tissue IF image; ICC validation is unreported (A06908-2 IF image caption and catalog applications). For work across human, mouse, and rat tissues, A06908-2 has IHC images in all three species; M06908-2 is a rabbit monoclonal, clone 21E91, with listed IHC and three-species reactivity but no IHC image in the payload (A06908-2 IHC image captions; M06908-2 catalog clone, applications, reactivity, and image alts).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry O75356 (ENTP5_HUMAN, Nucleoside diphosphate phosphatase ENTPD5).
- Human Protein Atlas. ENTPD5 tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. ENTPD5 subcellular location (ICC-IF): Secreted.
- Human Protein Atlas. ENTPD5 antibody validation summary (1 antibodies).
- PTEN phosphatase inhibits metastasis by negatively regulating the Entpd5/IGF1R pathway through ATF6. iScience 2023 — PMC9942123.
- Pten-null tumors cohabiting the same lung display differential AKT activation and sensitivity to dietary restriction. Cancer discovery 2013 — PMC3743121.
- ENTPD5 induces apoptosis in lung cancer cells via regulating caspase 3 expression. PloS one 2015 — PMC4368616.
- The UDPase ENTPD5 regulates ER stress-associated renal injury by mediating protein N-glycosylation. Cell death & disease 2023 — PMC9971188.
- PubMed PMID:9676430 — UniProt-cited evidence.
- PubMed PMID:10708485 — UniProt-cited evidence.
- PubMed PMID:15698960 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for ENTPD5 (Nucleoside diphosphate phosphatase ENTPD5). Expected localisation Cytoplasmic tissue staining (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A06908-2). Validated IHC antibody: A06908-2 at 2-5 μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Appendix (Endocrine cells); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human lung cancer tissue; fixative not specified (datasheet A06908-2); 2. Fixation: Image fixative and duration unreported (datasheet A06908-2); 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 A06908-2); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A06908-2); 8. Primary antibody: Rabbit anti-ENTPD5, 2-5 μg/ml (datasheet A06908-2); 9. Primary incubation: Overnight at 4 °C (datasheet A06908-2). Watch for: Fixation: Keep fixation consistent across sections (standard IHC practice; not target-specific); Caveat: A secreted variant may shift tissue staining (HPA tissue IHC); Isoform / epitope: No isoforms annotated; signal peptide 1–24 is cleaved (UniProt); Regulation: No expression regulator annotated (UniProt). Evidence: UniProt O75356, HPA tissue IHC · Enhanced, HPA ICC-IF · Secreted, Datasheet A06908-2, PMC3743121, PMC4368616. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/ENTPD5/immunohistochemistry-ihc-guide.