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NAMPT / Nicotinamide phosphoribosyltransferase · IHC design guide
Design Immunohistochemistry for NAMPT
Plan NAMPT staining in paraffin sections using the catalog antibody’s 0.5–1 μg/mL IHC range (datasheet PB10009). Assess the cytoplasmic tissue pattern while accounting for plasma positivity and secreted NAMPT (HPA tissue IHC; UniProt).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
NAMPT 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); nuclear location annotated (UniProt) | |
| Staining pattern | Cytoplasmic staining, highest in lymphoid and GI tissues (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet PB10009) | |
| Positive control | Adrenal gland+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 tissue samples (standard IHC practice; not target-specific) | |
| Caveat | Secreted NAMPT and plasma positivity complicate cell attribution (UniProt; HPA tissue IHC) | |
| Regulation | IL-1β stimulates endothelial secretion (UniProt) | |
| Isoform / epitope | No annotated isoforms; one 1–491 chain (UniProt) |
Section 1
Recommended NAMPT IHC & IF Protocols
The catalog antibody’s IHC-P protocol is complemented by 4 published NAMPT tissue-staining protocols (PMC7771344; PMC7771308; PMC7771238; PMC6411120).
| Sample | Paraffin-embedded mouse intestine tissue; fixative not specified (datasheet PB10009) |
| Fixation | Image fixative and duration unreported (datasheet PB10009); 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 PB10009); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet PB10009) |
| Primary antibody | Rabbit anti-NAMPT, 0.5-1μg/ml (datasheet PB10009) |
| Primary incubation | Overnight at 4 °C (datasheet PB10009) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet PB10009) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | NAMPT-positive staining in glandular cells of adrenal gland (HPA tissue IHC: Medium). HPA tissue profile: General cytoplasmic expression, highest abundance in lymphoid tissues and gastrointestinal tract. Positivity in plasma. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA pH 8.0 retrieval for the catalog antibody (datasheet: PB10009); published retrieval conditions apply to the antibodies used in those studies.
Section 2
What Is the Expected NAMPT Staining Pattern?
In paraffin section IHC, expect mainly cytoplasmic NAMPT in glandular cells, with strongest overall abundance in lymphoid tissues and the gastrointestinal tract (HPA tissue IHC). Nuclear staining can also be plausible, and extracellular signal may reflect secreted NAMPT (UniProt P43490). NAMPT has no transmembrane segment (UniProt P43490 topology). HPA rates its tissue pattern Approved, with external verification pending (HPA tissue IHC).
| Cytoplasmic staining in colon or duodenal glandular cells, around a medium level (HPA tissue IHC). | This matches the reported cell type, compartment, and level in these tissues (HPA tissue IHC). Compare cells within the section; a strong signal in every structure is less convincing than a cell-specific pattern (general IHC practice). |
| Granular nuclear staining alongside cytoplasmic signal. | Nuclear localisation is documented, including a predominantly granular nuclear pattern under non-inflammatory conditions (UniProt P43490). Treat staining confined to an unexpected compartment as a reason to review specificity and controls, rather than automatically rejecting nuclear signal (general IHC practice). |
| Strong staining in adipocytes or cardiomyocytes, which HPA lists as not detected (HPA tissue IHC). | This conflicts with those cell-level IHC observations, although it does not prove cross-reactivity (HPA tissue IHC). Check the no-primary control for endogenous detection activity and compare the cell pattern with an independently validated reagent (general IHC practice). |
| Uniform colour across cells, stroma, and empty spaces. | A uniform deposit does not resemble HPA's general cytoplasmic, cell-associated tissue pattern (HPA tissue IHC). Background from detection or nonspecific binding is possible; assess it with no-primary and appropriate negative controls (general IHC practice). |
| No signal in colon or duodenal glandular cells despite interpretable tissue (HPA tissue IHC). | These are reported medium-level positives, so absent staining warrants a workflow check (HPA tissue IHC). Verify retrieval, primary incubation, and detection against the antibody's validated IHC-P instructions; tissue positivity alone cannot identify which step failed (general IHC practice). |
💡Expected NAMPT appearanceA convincing positive is predominantly cytoplasmic staining in expected cells, such as medium-level colon or duodenal glandular cells (HPA tissue IHC); plausible nuclear signal may coexist (UniProt P43490), whereas uniform staining of unrelated structures calls for control review (general IHC practice).
| Tissue and cell selection | HPA reports general cytoplasmic expression, with highest abundance in lymphoid tissues and the gastrointestinal tract; its named glandular examples include medium-level colon and duodenum (HPA tissue IHC). Score the named cell population rather than treating a tissue-wide average as a cell-level result (general IHC practice). |
| Secreted NAMPT | HPA reports plasma positivity and cautions that a secreted protein variant can make RNA and protein locations differ (HPA tissue IHC). UniProt also lists NAMPT as secreted (UniProt P43490). Extracellular colour therefore needs anatomical and control context before it is scored as cellular staining (general IHC practice). |
| Antibody evidence | Two listed antibodies have Approved IHC status, HPA047776 and CAB034349 (HPA antibodies). The supplied HPA tissue profile is Approved but pending external verification (HPA tissue IHC); those labels support a starting expectation, not proof that every positive structure contains NAMPT. |
| Retrieval and epitope access | 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: Should nuclear speckles be expected in paraffin IHC? | A: HPA reports approved nuclear speckles and additional cell junction localisation in ICC-IF (HPA subcellular ICC-IF). Those are IF observations; the tissue IHC profile describes general cytoplasmic expression (HPA tissue IHC). Resolve each application against its own controls and images (general IHC practice). |
| Situation | Likely cause | Next action |
|---|---|---|
| Expected glandular cells have no visible signal. | A procedural failure or insufficient sensitivity is possible; colon and duodenal glandular cells are reported at medium level (HPA tissue IHC). | Check a known-positive section and no-primary control, then review retrieval, antibody incubation, and chromogen steps against the selected IHC-P instructions (general IHC practice). |
| Adipocytes appear strongly positive. | HPA reports adipocytes as not detected, so the observation needs verification; nonspecific binding or endogenous detection activity are possible (HPA tissue IHC; general IHC practice). | Compare adjacent morphology and a no-primary control; reassess the result with an independently validated IHC antibody if available (general IHC practice). |
| Nearly every compartment shows the same brown deposit. | This does not fit HPA's general cytoplasmic tissue profile and may reflect detection background (HPA tissue IHC; general IHC practice). | Inspect no-primary and negative controls, then review blocking, washes, and chromogen development as general IHC steps (general IHC practice). |
| Only nuclear staining is seen. | Nuclear NAMPT is documented, but the dominant tissue IHC pattern is cytoplasmic (UniProt P43490; HPA tissue IHC). | Check whether signal follows nuclei consistently in the expected cells; compare with the IHC antibody's validation images and controls before scoring it (general IHC practice). |
| Plasma or extracellular material stains strongly. | Plasma positivity is reported, and NAMPT can be secreted (HPA tissue IHC; UniProt P43490). | Record the extracellular compartment separately from cell staining and inspect controls before calling the signal specific (general IHC practice). |
| A positive section is much darker than the reported medium-level glandular examples. | Those HPA levels describe particular cell types, not a universal intensity ceiling; overdevelopment or nonspecific signal also remains possible (HPA tissue IHC; general IHC practice). | Compare the same cell type and tissue with controls, then review primary dilution and chromogen time under the selected IHC-P instructions (general IHC practice). |
Experiment evidence
IHC & IF images showing NAMPT staining
PB10009 · IHCIHC analysis of Visfatin using anti-Visfatin antibody (PB10009). Visfatin was detected in a paraffin-embedded section of mouse intestine tis
PB10009 · IHCIHC analysis of Visfatin using anti-Visfatin antibody (PB10009). Visfatin was detected in a paraffin-embedded section of human mammary cance
PB10009 · IFIF analysis of Visfatin/NAMPT using anti-Visfatin/NAMPT antibody (PB10009). Visfatin/NAMPT was detected in an immunocytochemical section of
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000105835-NAMPT/tissue/cerebral+cortex#imid_13987756
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000105835-NAMPT/tissue/colon#imid_13987664
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000105835-NAMPT/tissue/liver#imid_13987673
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000105835-NAMPT/tissue/kidney#imid_13987682
HPA · ICC-IF A-549Human Protein Atlas immunofluorescence, A-549, antibody HPA057722; green = NAMPT, blue = nucleus, red = microtubules.
HPA · ICC-IF A-549Human Protein Atlas immunofluorescence, A-549, antibody HPA057722; green = NAMPT, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for NAMPT IHC & IF
🧪Run adrenal gland first and look for staining in glandular cells (Medium; HPA: adrenal gland, glandular cells). Use adipose tissue adipocytes as the biological tissue negative (HPA: adipocytes not detected); on the adrenal slide, assess background in neighboring cells that lack specific staining, without assuming any named cell population is NAMPT-negative.
Positive control tissue: Adrenal gland (Glandular cells, HPA Medium)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show NAMPT in A-549, HEK293, U2OS, with annotated localisation: Nuclear speckles (approved) (HPA subcellular).
Technical controls: Include a no-primary (secondary-only) slide and a concentration-matched, nonimmune rabbit IgG isotype control for the rabbit catalog antibody (selected-SKU IHC caption: rabbit primary). Confirm specificity with NAMPT-knockout material or a validated peptide-block experiment; with the caption’s biotin-based DAB detection, check endogenous peroxidase and biotin background on the positive tissue (selected-SKU IHC caption: biotinylated secondary, SABC, DAB).
⚠️Feasibility: A NAMPT-specific fixation window or fixation effect is unreported, and the selected-SKU paraffin-section caption does not state a fixative (selected-SKU IHC caption: fixative not stated). Heat retrieval in EDTA at pH 8.0 is documented for that IHC example, but retrieval dependence and whether frozen sections or IF are easier are unreported (selected-SKU IHC caption: heat-mediated EDTA retrieval). For adrenal gland, inspect glandular and neighboring cells separately for detection background; the supplied HPA row identifies glandular staining but reports no tissue-specific artefact (HPA: adrenal gland, glandular cells Medium).
HPA tissue IHC evidence for NAMPT
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — At least one protein variant secreted, tissue location of RNA and protein might differ and correlation is complex. Pending external verification.). 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 | Medium | Protein (IHC) | HPA → |
| Appendix | Glandular cells | Medium | Protein (IHC) | HPA → |
| Breast | Glandular cells | Medium | Protein (IHC) | HPA → |
| Cervix | Glandular cells | Medium | Protein (IHC) | HPA → |
| Colon | 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 → |
| Caudate | Glial cells | Not detected | Protein (IHC) | HPA → |
| Cerebellum | Cells in granular layer | Not detected | Protein (IHC) | HPA → |
| Heart muscle | Cardiomyocytes | Not detected | Protein (IHC) | HPA → |
| Hippocampus | Glial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced NAMPT IHC Tips
Troubleshoot NAMPT staining in paraffin sections by checking retrieval, cellular compartment, detection background and cell-specific scoring (datasheet PB10009; HPA tissue IHC).
How should I retrieve NAMPT antigen when staining is weak in paraffin sections?
Start with heat-mediated retrieval in EDTA at pH 8.0 (datasheet PB10009). The selected paraffin-section example used this retrieval before applying 1 μg/ml primary antibody overnight at 4°C (datasheet PB10009). If signal is weak, check heating consistency, section adhesion and antibody incubation against a concurrently processed positive section before changing the retrieval condition (standard IHC practice). Compare retrieval changes on adjacent sections with identical detection and DAB development, since stronger brown staining can also reflect increased background (standard IHC practice). Record nuclear and cytoplasmic staining separately when assessing improvement (UniProt P43490 subcellular location; HPA tissue IHC).
Could fixation explain inconsistent NAMPT staining across paraffin blocks?
Target-specific fixation sensitivity is unknown: the selected paraffin-section caption does not state a fixative (datasheet PB10009). Record the fixative, fixation interval and processing history for each block before comparing staining intensity (standard IHC practice). Process adjacent sections together with the same EDTA pH 8.0 retrieval and detection conditions to test whether the difference persists (datasheet PB10009; standard IHC practice). Include a concurrently stained tissue control and a no-primary control to distinguish lost antigen signal from processing or detection background (standard IHC practice). Do not assign a NAMPT-specific fixation effect from its tissue pattern, intracellular location or modified residues (HPA tissue IHC; UniProt P43490 subcellular location and modified residues).
Should NAMPT staining be nuclear, cytoplasmic or extracellular in tissue?
Evaluate cytoplasmic staining because the tissue IHC profile describes general cytoplasmic expression, with highest abundance in lymphoid tissues and the gastrointestinal tract (HPA tissue IHC). Nuclear staining is also biologically plausible: NAMPT is annotated in the nucleus, and a granular nuclear pattern is described under non-inflammatory conditions (UniProt P43490 subcellular location). Nuclear speckles and cell junctions are reported from cell-based imaging; verify any corresponding tissue pattern with controls before treating it as established IHC localisation (HPA subcellular; standard IHC practice). Plasma positivity is reported, but extracellular staining alone cannot identify its cellular source (HPA tissue IHC; standard IHC interpretation). Score compartments separately and check morphology on the counterstained section (standard IHC practice).
Could isoforms or epitope accessibility explain discordant NAMPT staining?
The supplied record lists 0 isoforms and one chain spanning residues 1–491, so an isoform-specific interpretation is unsupported here (UniProt P43490 processing and isoforms). NAMPT has no annotated transmembrane segment or signal peptide, consistent with evaluating intracellular antigen access in sections (UniProt P43490 topology and processing). Its modified residues include phosphotyrosine 188 and phosphoserine 472, but the supplied caption does not map the antibody epitope (UniProt P43490 modified residues; datasheet PB10009). Do not attribute discordant staining to either modification without epitope or perturbation evidence (standard IHC interpretation). Recheck retrieval and controls across sections before proposing molecular variants as the cause (standard IHC practice).
How can IF help resolve ambiguous chromogenic NAMPT localisation?
Use IF as a complementary localisation check for the paraffin-section IHC result, keeping the IHC-validated antibody’s tissue performance as the reference (datasheet PB10009; standard IHC practice). Multiplex NAMPT with a marker for the expected cell population, such as a glandular-cell or macrophage marker chosen for the specimen, and inspect each channel independently (HPA tissue IHC; standard IF practice). Choose spectrally separated fluorophores and assess unstained tissue autofluorescence before assigning a faint signal to NAMPT (standard IF practice). Because NAMPT has no transmembrane segment and is annotated in nuclear and cytoplasmic compartments, permeabilisation must allow access to intracellular epitopes (UniProt P43490 topology and subcellular location; standard IF practice). Confirm apparent colocalisation against morphology and single-stain controls (standard IF practice).
What causes diffuse brown background in NAMPT chromogenic IHC?
Separate diffuse DAB deposition from cellular staining by examining a no-primary section processed through the same detection reagents (standard IHC practice). The selected example used 10% goat serum block, biotinylated goat anti-rabbit secondary antibody, streptavidin-biotin complex and DAB (datasheet PB10009). Apply a peroxidase block and assess endogenous biotin where relevant to this detection chemistry; those are general workflow controls, not NAMPT-specific findings (standard IHC practice). Compare background after thorough washes and matched DAB development, then adjust blocking or detection conditions if the no-primary section remains brown (standard IHC practice). Interpret staining beside tissue morphology because plasma positivity can coexist with cellular NAMPT expression (HPA tissue IHC).
How should I quantify NAMPT IHC when staining spans multiple compartments? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and region before scoring, then report nuclear and cytoplasmic staining separately (UniProt P43490 subcellular location; HPA tissue IHC; standard IHC practice). For cellular staining, record percent positive cells and intensity grades 0–3; an H-score combines these into a 0–300 range (standard IHC scoring practice). For infiltrating positive cells, report density per mm² of viable tissue and identify the counted compartment (standard IHC quantification practice). Normalise comparisons to the same cell class, sampled tissue area, staining batch and exposure or scoring rules, with matched control sections (standard IHC practice). Keep plasma-associated signal outside a cellular H-score unless a separate extracellular measurement is defined (HPA tissue IHC; standard IHC quantification practice).
How can I distinguish true NAMPT staining from tissue or detection artefact?
Look for reproducible cellular signal in a plausible compartment: tissue IHC commonly shows cytoplasmic NAMPT, while the protein is also annotated in the nucleus (HPA tissue IHC; UniProt P43490 subcellular location). Check cell identity against morphology; reported gastrointestinal glandular-cell positivity and lung macrophage staining offer context for those respective specimens (HPA tissue IHC). Treat staining confined to section edges, folds or necrotic areas as suspect until it persists in intact tissue and repeat sections (standard IHC practice). A no-primary control helps identify endogenous peroxidase or detection background in DAB sections (standard IHC practice). Plasma positivity and a secreted form make extracellular signal possible, but extracellular brown deposit alone does not establish its source (HPA tissue IHC; UniProt P43490 subcellular location).
Boster reagents
Best NAMPT / Nicotinamide phosphoribosyltransferase IHC Antibodies
PB10009 has real IHC images from paraffin sections of human mammary cancer, mouse intestine, and rat intestine (PB10009 IHC captions), plus IF data from A549 cells (PB10009 IF caption).
Real IHC data
Anti-Visfatin/NAMPT Antibody ®
Cat # PB10009
PB10009 is listed for IHC, IHC-F, IF, and ICC, with Human, Mouse, and Rat reactivity (catalog: PB10009). Its IHC captions show paraffin sections of human mammary cancer, mouse intestine, and rat intestine; its IF caption shows A549 cells (PB10009 image captions).
Which to pick: Choose PB10009 for paraffin-section IHC in the pictured tissues and species, using its 1 μg/ml primary-antibody image conditions as a starting point (PB10009 IHC captions); the fixative is unreported (PB10009 IHC captions). For IF/ICC, PB10009 is the listed option and has an A549 cell IF image (catalog: PB10009; PB10009 IF caption). For cross-species IHC, PB10009 has pictured human, mouse, and rat sections (PB10009 IHC captions); clonality is unreported.
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P43490 (NAMPT_HUMAN, Nicotinamide phosphoribosyltransferase).
- Human Protein Atlas. NAMPT tissue IHC expression (reliability: Approved).
- Human Protein Atlas. NAMPT subcellular location (ICC-IF): Mainly localized to the nuclear speckles. In addition localized to the cell junctions..
- Human Protein Atlas. NAMPT antibody validation summary (3 antibodies).
- Nicotinamide Phosphoribosyl Transferase a Reliable Marker of Progression in Cervical Dysplasia. Anticancer research 2017 — PMC7771344.
- Nicotinamide Phosphoribosyl Transferase Is Increased in Osteosarcomas and Chondrosarcomas Compared to Benign Bone and Cartilage. Anticancer research 2019 — PMC7771308.
- Cystathione β-Synthase Is Increased in Thyroid Malignancies. Anticancer research 2018 — PMC7771238.
- Nicotinamide phosphoribosyltransferase expression and clinical outcome of resected stage I/II pancreatic ductal adenocarcinoma. PloS one 2019 — PMC6411120.
- PubMed PMID:8289818 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
- PubMed PMID:12853948 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for NAMPT (Nicotinamide phosphoribosyltransferase). Expected localisation Cytoplasmic tissue staining (HPA tissue IHC); nuclear location annotated (UniProt). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet PB10009). Validated IHC antibody: PB10009 at 0.5-1μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Glandular cells); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded mouse intestine tissue; fixative not specified (datasheet PB10009); 2. Fixation: Image fixative and duration unreported (datasheet PB10009); 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 PB10009); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet PB10009); 8. Primary antibody: Rabbit anti-NAMPT, 0.5-1μg/ml (datasheet PB10009); 9. Primary incubation: Overnight at 4 °C (datasheet PB10009). Watch for: Fixation: Keep fixation consistent across tissue samples (standard IHC practice; not target-specific); Caveat: Secreted NAMPT and plasma positivity complicate cell attribution (UniProt; HPA tissue IHC); Isoform / epitope: No annotated isoforms; one 1–491 chain (UniProt); Regulation: IL-1β stimulates endothelial secretion (UniProt). Evidence: UniProt P43490, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the nuclear speckles. In addition localized to…, Datasheet PB10009, PMC7771344, PMC7771308. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/NAMPT/immunohistochemistry-ihc-guide.