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NNMT / Nicotinamide N-methyltransferase · IHC design guide
Design Immunohistochemistry for NNMT
Plan NNMT IHC-P around cytoplasmic staining in hepatocytes and kidney tubule cells (HPA tissue IHC). The catalog antibody has an IHC-P dilution of 1:10–1:50 (datasheet: A02676-1 IHC-P); compare sections with consistent fixation and appropriate controls (standard IHC practice).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
NNMT 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 in a few tissues (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic signal in hepatocytes and kidney tubule cells (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Kidney+4 more · see all | |
| Negative control | Adipose tissue+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Formalin-fixed paraffin liver is shown (selected-SKU IHC image A02676-1); Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A02676-1) | |
| Caveat | Antibody staining has low consistency with RNA expression (HPA tissue IHC) | |
| Regulation | Predominantly expressed in liver (UniProt) | |
| Isoform / epitope | 0 annotated isoforms or cleavage events; no mapped epitope shift (UniProt) |
Section 1
Recommended NNMT IHC & IF Protocols
The catalog antibody’s IHC-P protocol is accompanied by published NNMT staining methods for prostate tissue, ovarian clear cell carcinoma, and paraffin sections (PMC4114624; PMC12378295; PMC12098763).
| Sample | FFPE human liver tissue (datasheet A02676-1) |
| Fixation | Image formalin-fixed; duration unreported (datasheet A02676-1); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat-induced epitope retrieval in citrate buffer, pH 6.0, 20 min at 95–98 °C (standard rule: cytoplasmic / membrane antigen) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% normal serum of the secondary host, 30 min, room temperature (standard) |
| Primary antibody | Rabbit anti-NNMT, 1:10-1:50 (datasheet A02676-1) |
| Primary incubation | Overnight at 4 °C (standard) |
| Detection | HRP-polymer secondary, DAB chromogen 5–10 min (standard) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | NNMT-positive staining in cells in tubules of kidney (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in a few tissues. No signal in the no-primary control. |
💡Decision noteStart with citrate pH 6.0 HIER at 95–98 °C for 20 min (page retrieval); use each article’s stated retrieval conditions when reproducing its method.
Section 2
What Is the Expected NNMT Staining Pattern?
NNMT is a cytoplasmic protein without a transmembrane segment (UniProt P40261). In paraffin-section IHC, expect strong cytoplasmic staining in hepatocytes, kidney tubular cells and pancreatic exocrine glandular cells (HPA tissue IHC: High). Interpret tissue comparisons cautiously: HPA rates its antibody staining Approved but reports low consistency with RNA expression (HPA tissue IHC: reliability).
| Strong cytoplasmic signal in hepatocytes or kidney tubular cells, with cellular boundaries still readable. | This fits the reported compartment and high-staining cell populations (UniProt P40261: Cytoplasm; HPA tissue IHC: High in hepatocytes and kidney tubular cells). Judge the cells stained as well as the overall intensity; a positive-looking region alone does not establish the expected pattern (general IHC practice). |
| Predominantly nuclear, membranous or extracellular signal in an otherwise positive-looking area. | These compartments do not match the reported cytoplasmic localization or lack of a transmembrane segment (UniProt P40261). Review morphology and controls before assigning NNMT positivity; misplaced chromogen can reflect nonspecific staining or detection artefact (general IHC practice). |
| Strong staining chiefly in adipocytes or hematopoietic cells. | HPA reports NNMT as not detected in those cell populations (HPA tissue IHC: adipocytes; bone marrow hematopoietic cells). Consider cross-reactivity or endogenous detection activity, then inspect matched controls (general IHC practice). A reported negative population is a comparison, not proof that every specimen must be negative. |
| Diffuse chromogen across several tissue compartments, including spaces without recognizable cells. | That distribution does not resemble the reported cell-associated cytoplasmic pattern (HPA tissue IHC: cytoplasmic expression in a few tissues). Check the primary-omitted control and background from the detection system; widespread haze makes intensity scoring unreliable (general IHC practice). |
| No convincing signal in hepatocytes on a liver section. | HPA reports high hepatocyte staining and liver-enriched RNA, while UniProt describes predominant liver expression (HPA tissue IHC; UniProt P40261: tissue specificity). First confirm tissue morphology and control performance, then review the IHC-validated antibody's documented conditions (general IHC practice). |
💡Expected NNMT appearanceCall a result consistent when hepatocytes or kidney tubular cells show strong cytoplasmic staining (HPA tissue IHC: High; UniProt P40261: Cytoplasm); predominant nuclear or extracellular signal, or widespread acellular chromogen, argues for artefact or nonspecific detection (general IHC practice).
| Tissue and cell selection | Hepatocytes, kidney tubular cells and pancreatic exocrine glandular cells are reported High; cervix, endometrium and epididymis glandular cells are Medium (HPA tissue IHC). Compare the named cell populations, since adjacent cells need not share the same score (general IHC practice). |
| Tissue evidence conflict | Pancreatic exocrine cells stain High by HPA IHC, whereas UniProt says NNMT was not detected in pancreas (HPA tissue IHC; UniProt P40261: tissue specificity). Treat pancreas as an observed IHC pattern with this explicit caveat, rather than a settled expression control. |
| Antibody validation | HPA059180 is Approved for IHC, but HPA reports low consistency between antibody staining and RNA expression (HPA antibodies; HPA tissue IHC: reliability). The supplied status is Approved, so do not infer an Enhanced validation claim from it. |
| Topology and processing | NNMT is cytoplasmic, lacks a transmembrane segment and signal peptide, and is annotated as one chain spanning residues 1–264 (UniProt P40261). These annotations support a cellular cytoplasmic expectation; they do not identify an antibody epitope or predict antigen retrieval performance. |
| IF/ICC question: should Golgi staining match tissue IHC? | HPA ICC-IF places NNMT mainly at the Golgi apparatus and additionally in the cytosol, whereas its tissue IHC profile is cytoplasmic (HPA subcellular; HPA tissue IHC). Compare each observation with its own assay and specimen context; the IF/ICC workflow belongs to its separate guide. |
| Situation | Likely cause | Next action |
|---|---|---|
| Liver section has no hepatocyte signal. | This conflicts with the reported High hepatocyte pattern (HPA tissue IHC); the image alone does not identify the technical cause. | Confirm the expected cells are present, inspect run controls, then review the catalog antibody's IHC-P instructions for retrieval, dilution and detection conditions (general IHC practice). Do not infer a target-specific fixation effect from this result. |
| Kidney staining is scored from the whole section rather than tubules. | The reported High signal is assigned specifically to cells in tubules (HPA tissue IHC), so a region-wide score can obscure the relevant cell population. | Recheck morphology and score tubular cells separately from other compartments (general IHC practice). Compare localization with the cytoplasmic expectation (UniProt P40261). |
| Pancreatic exocrine cells stain strongly, but the result seems inconsistent with the expression summary. | HPA reports High exocrine-cell IHC staining, while UniProt reports no pancreatic detection (HPA tissue IHC; UniProt P40261: tissue specificity). | Record both observations and examine antibody and detection controls before interpreting the signal (general IHC practice). Avoid using pancreas alone to settle the discrepancy. |
| Adipocytes, hematopoietic cells or acellular areas dominate the positive signal. | The named cell populations are reported Not detected (HPA tissue IHC); acellular staining also departs from a cellular cytoplasmic pattern (HPA tissue IHC). | Examine a primary-omitted control and detection background, then reassess whether signal follows cell morphology (general IHC practice). Treat unexplained staining as unresolved specificity, not confirmed NNMT. |
| Signal is predominantly nuclear or outlines cell membranes. | That localization differs from cytoplasm, and NNMT has no annotated transmembrane segment (UniProt P40261). | Check section morphology, counterstain and control slides; adjust interpretation only after confirming where the chromogen sits within cells (general IHC practice). Do not score compartment-mismatched signal as an expected positive. |
| Tissue IHC looks diffuse while a separate IF/ICC image appears Golgi enriched. | HPA describes cytoplasmic tissue IHC and mainly Golgi, additionally cytosolic, ICC-IF localization (HPA tissue IHC; HPA subcellular). | Document the assay and specimen for each image, then compare each with its corresponding HPA pattern. If the IHC signal lacks recognizable cellular localization, revisit IHC controls before scoring (general IHC practice). |
Experiment evidence
IHC & IF images showing NNMT staining
A02676-1 · IHCNNMT Antibody (Center) (Cat. #A02676-1)immunohistochemistry analysis in formalin fixed and paraffin embedded human liver tissue followed by
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000166741-NNMT/tissue/cerebral+cortex#imid_16368395
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000166741-NNMT/tissue/colon#imid_16364657
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000166741-NNMT/tissue/liver#imid_16364658
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000166741-NNMT/tissue/kidney#imid_16364651
HPA · ICC-IF RPTEC/TERT1Human Protein Atlas immunofluorescence, RPTEC/TERT1, antibody HPA059180; green = NNMT, blue = nucleus, red = microtubules.
HPA · ICC-IF RPTEC/TERT1Human Protein Atlas immunofluorescence, RPTEC/TERT1, antibody HPA059180; green = NNMT, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for NNMT IHC & IF
🧪Run liver first: hepatocytes should stain (HPA: High in hepatocytes; selected A02676-1 caption: liver IHC). Use adipose tissue as the negative tissue: adipocytes should show no specific staining (HPA: Not detected in adipocytes); on the liver slide, cells without hepatocyte staining should remain at background, but no other liver cell type is established as negative by the supplied HPA rows.
Positive control tissue: Kidney (Cells in tubules, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show NNMT in RPTEC/TERT1, Rh30, SiHa, with annotated localisation: Golgi apparatus (approved) (HPA subcellular).
Technical controls: Include a no-primary (secondary-only) control, an isotype control matched to the primary antibody’s host species and clonality, and a biological specificity control using NNMT knockout material or peptide blocking (standard IHC practice). Quench endogenous peroxidase for the liver HRP/DAB assay, and block endogenous biotin if using avidin-biotin detection (selected A02676-1 caption: peroxidase/DAB; standard IHC practice).
⚠️Feasibility: The selected A02676-1 liver caption explicitly reports formalin-fixed, paraffin-embedded tissue, but gives no target-specific fixation window (selected A02676-1 caption). Retrieval dependence is unreported; the supplied evidence also does not establish whether frozen sections or IF are easier. Endogenous biotin in liver can cause background with avidin-biotin detection (standard IHC practice).
HPA tissue IHC evidence for NNMT
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Low 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 |
|---|---|---|---|---|
| Kidney | Cells in tubules | High | Protein (IHC) | HPA → |
| Liver | Hepatocytes | High | Protein (IHC) | HPA → |
| Pancreas | Exocrine glandular cells | High | Protein (IHC) | HPA → |
| Cervix | Glandular cells | Medium | Protein (IHC) | HPA → |
| Endometrium | 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 → |
| Appendix | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Breast | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced NNMT IHC Tips
These NNMT troubleshooting questions focus on chromogenic IHC in paraffin sections, with one entry for IF/ICC.
How should I retrieve NNMT in paraffin sections when staining is weak?
Start with citrate buffer at pH 6.0, heated to 95–98 °C for 20 min (page retrieval setting). Let sections cool consistently, then compare the same IHC-validated antibody on a liver control and the test specimen in one staining run (UniProt P40261: liver-predominant expression; HPA: high in hepatocytes). If staining remains weak, pilot a different retrieval buffer or pH on adjacent sections while keeping antibody dilution and detection conditions fixed (IHC optimisation practice). Record both signal and tissue damage, because stronger colour alone does not establish specific staining (IHC interpretation practice).
Can I adjust fixation to recover weak NNMT staining?
The selected antibody has been shown on formalin-fixed, paraffin-embedded human liver with peroxidase-linked secondary detection and DAB (A02676-1 tissue-IHC caption). That image does not establish how sensitive NNMT staining is to fixation duration or fixative choice (A02676-1 tissue-IHC caption). If fixation varies among samples, compare sections with documented processing histories using the same retrieval and staining run, and include a liver control (IHC optimisation practice; UniProt P40261: liver-predominant expression). Interpret a weak result cautiously until processing differences and tissue preservation have been assessed, rather than assigning it to a target-specific fixation effect (IHC interpretation practice).
Should NNMT stain the cytoplasm, Golgi, or cell surface in IHC?
Expect intracellular staining: UniProt places NNMT in the cytoplasm, while HPA subcellular imaging reports mainly Golgi localisation with additional cytosol (UniProt P40261: cytoplasm; HPA subcellular: Golgi and cytosol). NNMT has no annotated transmembrane segment or signal peptide, so crisp cell-surface staining warrants closer scrutiny (UniProt P40261: topology and processing). In paraffin liver sections, assess the staining within hepatocytes alongside their morphology, rather than requiring a Golgi pattern resolvable by light microscopy (HPA tissue IHC: high in hepatocytes; IHC interpretation practice). Check whether apparent membrane staining persists after reducing detection intensity and reviewing an appropriate negative control (IHC troubleshooting practice).
Could an NNMT isoform or modified epitope explain inconsistent staining?
The supplied UniProt record lists one 264-amino-acid chain and no annotated isoforms or domains, so an isoform-specific explanation lacks support here (UniProt P40261: processing, isoforms and domains). It does list modified residues, including acetyllysine at position 39 and citrullination annotations, but supplies no antibody epitope map (UniProt P40261: modified residues). Do not attribute lost staining to any one modification without direct epitope evidence (UniProt P40261: modified residues; antibody evidence supplied here). Compare adjacent sections across retrieval conditions and verify the antibody’s documented IHC application before interpreting a regional difference as biology (IHC optimisation practice).
How can I assess NNMT by multiplex IF in tissue?
For IF/ICC, pair NNMT with an independently validated marker for the expected cell type, such as hepatocytes in liver, and examine channel overlap at the cell level (HPA tissue IHC: high in hepatocytes; IF practice). Inspect an unstained section for autofluorescence before choosing fluorophores, and favour a well-separated far-red channel when shorter wavelengths have substantial background (IF practice). Apply controlled permeabilisation for intracellular epitope access: NNMT has no annotated transmembrane segment, although the antibody’s epitope is unspecified (UniProt P40261: topology; IF practice). Include single-colour and secondary-only controls to distinguish signal, bleed-through and background; assess Golgi-like and cytosolic patterns separately (IF practice; HPA subcellular: Golgi and cytosol).
What should I check when NNMT DAB staining is diffuse or widespread?
First inspect a section processed without primary antibody to identify detection-system background, and compare it with a liver control stained in the same run (IHC practice; UniProt P40261: liver-predominant expression). For peroxidase-based DAB detection, include an endogenous peroxidase block and control the DAB development time across sections (IHC practice; A02676-1 tissue-IHC caption: peroxidase-linked secondary and DAB). Review blocking, washes and antibody concentration using the validated protocol, then change one variable at a time (IHC optimisation practice). Diffuse staining outside intact cells deserves particular caution because reported NNMT tissue staining is cytoplasmic and limited to selected cell populations (HPA tissue IHC: cytoplasmic expression in a few tissues).
How should I score NNMT staining across paraffin tissue samples? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and viable tissue area before scoring; in liver, hepatocytes are a supported population to assess (HPA tissue IHC: high in hepatocytes). For chromogenic NNMT staining, record the percentage of positive target cells and an intensity score, then calculate an H-score from 0–300 if that method fits the study (IHC scoring practice). Alternatively, report positive-cell density per mm², normalised to viable tissue area, and keep segmentation and DAB thresholds fixed across the batch (IHC quantification practice). Report controls, processing differences and observer rules alongside scores, because HPA flags low consistency between antibody staining and RNA expression data (HPA tissue IHC: Approved reliability description).
How do I distinguish true NNMT staining from artefact?
A credible signal should occur within intact cells in a plausible intracellular compartment; liver hepatocytes and kidney tubule cells provide reported high-staining examples (UniProt P40261: cytoplasm; HPA tissue IHC: high in hepatocytes and tubule cells). Review unexpected cell-surface or purely nuclear colour against tissue morphology and negative controls, since NNMT lacks a transmembrane segment and is reported in cytoplasm, Golgi and cytosol (UniProt P40261: topology and localisation; HPA subcellular: Golgi and cytosol). Exclude section-edge colour, necrotic areas and residual endogenous peroxidase signal during scoring (IHC interpretation practice). Treat pancreatic staining cautiously: HPA reports high exocrine staining, whereas UniProt reports NNMT as undetected in pancreas (HPA tissue IHC: pancreas; UniProt P40261: tissue specificity).
Boster reagents
Best NNMT / Nicotinamide N-methyltransferase IHC Antibodies
The catalog shows human liver IHC data for A02676-1 (image caption) and human ICC/IF validation for A02676-2 (applications and reactivity); A02676-2 also lists mouse and rat reactivity (catalog reactivity).
Real IHC data
Anti-NNMT Antibody (Center)
Cat # A02676-1
The rendered A02676-1 card shows IHC-P staining of formalin-fixed, paraffin-embedded human liver with peroxidase/DAB detection (A02676-1 image caption). A02676-2 is listed for human ICC/IF, but has no IF figure in the supplied payload (A02676-2 applications, reactivity and image alts).
Which to pick: Choose A02676-1 for human tissue IHC-P: it is polyclonal, has a 1:10–1:50 IHC-P dilution, and its own image documents formalin-fixed, paraffin-embedded human liver (A02676-1 catalog and image caption). Choose A02676-2 for human ICC/IF at a listed 5 μg/ml; its applications do not include IHC-P (A02676-2 catalog). A02676-2 lists human, mouse and rat reactivity, but the supplied ICC/IF validation is human only (A02676-2 catalog applications, reactivity and dilution data).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P40261 (NNMT_HUMAN, Nicotinamide N-methyltransferase).
- Human Protein Atlas. NNMT tissue IHC expression (reliability: Approved).
- Human Protein Atlas. NNMT subcellular location (ICC-IF): Mainly localized to the Golgi apparatus. In addition localized to the cytosol..
- Human Protein Atlas. NNMT antibody validation summary (1 antibodies).
- Nicotinamide N-methyltransferase is overexpressed in prostate cancer and correlates with prolonged progression-free and overall survival times. Oncology letters 2014 — PMC4114624.
- Nicotinamide N-methyltransferase enhances paclitaxel resistance in ovarian clear cell carcinoma. Human cell 2025 — PMC12378295.
- NNMT/1-MNA protects against hepatic ischemia-reperfusion injury through the AKT/FOXO1/ANGPT2/JNK axis. Nature communications 2025 — PMC12098763.
- Targeting Nicotinamide N-Methyltransferase and miR-449a in EGFR-TKI-Resistant Non-Small-Cell Lung Cancer Cells. Molecular therapy. Nucleic acids 2018 — PMC5992482.
- PubMed PMID:8182091 — UniProt-cited evidence.
- PubMed PMID:8575745 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for NNMT (Nicotinamide N-methyltransferase). Expected localisation Cytoplasmic staining in a few tissues (HPA tissue IHC). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: A02676-1 at 1:10-1:50 (the product's own IHC validation image is shown). Controls (HPA): positive Kidney (Cells in tubules); negative Adipose tissue. IHC-P workflow on FFPE sections. Protocol steps: 1. Sample: FFPE human liver tissue (datasheet A02676-1); 2. Fixation: Image formalin-fixed; duration unreported (datasheet A02676-1); 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-induced epitope retrieval in citrate buffer, pH 6.0 , 20 min at 95–98 °C…; 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% normal serum of the secondary host, 30 min , room temperature (standard); 8. Primary antibody: Rabbit anti-NNMT, 1:10-1:50 (datasheet A02676-1); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Formalin-fixed paraffin liver is shown (selected-SKU IHC image A02676-1); Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A02676-1); Caveat: Antibody staining has low consistency with RNA expression (HPA tissue IHC); Isoform / epitope: 0 annotated isoforms or cleavage events; no mapped epitope shift (UniProt); Regulation: Predominantly expressed in liver (UniProt). Evidence: UniProt P40261, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the Golgi apparatus. In addition localized to…, Datasheet A02676-1, PMC4114624, PMC12378295. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/NNMT/immunohistochemistry-ihc-guide.