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TNFAIP8L3 / Tumor necrosis factor alpha-induced protein 8-like protein 3 · IHC design guide
Design Immunohistochemistry for TNFAIP8L3
Plan TNFAIP8L3 staining in paraffin sections using smooth muscle as a strong tissue reference (HPA tissue IHC). The human-reactive catalog antibody PA2257 lists 0.5–1 μg/mL for IHC-P (datasheet).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
TNFAIP8L3 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 and membranous tissue staining (HPA tissue IHC) | |
| Staining pattern | Strongest in smooth muscle, with membrane and cytoplasmic staining (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Bronchus+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 paraffin sections. (standard IHC practice; not target-specific) | |
| Caveat | PDGF activation can shift cytoplasmic signal to the membrane (UniProt) | |
| Regulation | Expression regulation unreported (UniProt) | |
| Isoform / epitope | No isoforms or processing annotated; epitope region unreported (UniProt) |
Section 1
Recommended TNFAIP8L3 IHC & IF Protocols
The catalog antibody’s IHC protocol is accompanied by published TNFAIP8L3 staining methods for lung adenocarcinoma and lung cancer tissue (PMC12404833; PMC5840720; PMC7947417).
| Sample | Tissue sections; selected-image fixative not specified (standard IHC workflow) |
| Fixation | Image fixative and duration unreported (datasheet PA2257); 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-TNFAIP8L3, 0.5-1μg/ml (datasheet PA2257) |
| 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 | TNFAIP8L3-positive staining in ciliated cells (ciliary rootlets) of bronchus (HPA tissue IHC: High). HPA tissue profile: Membrane and cytoplasmic expression in most tissues, most abundant in smooth muscle. No signal in the no-primary control. |
💡Decision noteStart with citrate pH 6.0 retrieval at 95–98 °C for 20 min (page retrieval); adjust if needed for the published method’s shorter boil (PMC12404833).
Section 2
What Is the Expected TNFAIP8L3 Staining Pattern?
TNFAIP8L3 is expected in the cytoplasm and at the cell membrane, with prominent staining in smooth muscle and high staining in several glandular cell populations (UniProt Q5GJ75; HPA: tissue IHC). It has no annotated transmembrane segment (UniProt Q5GJ75 topology). Treat this as a working pattern: HPA rates the tissue staining Approved, with external verification pending (HPA: tissue IHC reliability).
| Cytoplasmic staining, sometimes with membrane staining, appears in smooth muscle or glandular cells. | This fits the reported compartment pattern; smooth muscle is the most abundant site in the tissue profile, while colon, duodenum and endometrial glandular cells are scored High (HPA: tissue IHC). Compare like cell types within the section: the HPA levels describe observed staining, not a universal intensity threshold. |
| A paraffin section shows strong nuclear-only staining with little cytoplasmic or membrane signal. | Flag the compartment mismatch and review the control and detection steps before assigning it to TNFAIP8L3 (HPA: tissue IHC; UniProt Q5GJ75 localization). Nuclear-only staining is not the reported tissue IHC pattern. HPA separately reports an additional nucleoplasmic location in ICC-IF, so the two assay observations should be interpreted in their own contexts (HPA: subcellular ICC-IF). |
| Prominent staining appears in adipocytes or cardiomyocytes. | Both cell populations are scored Not detected in the supplied tissue images (HPA: adipose tissue adipocytes; HPA: heart muscle cardiomyocytes). Consider cross-reactivity or endogenous chromogenic activity, especially if the same signal appears in a no-primary control (general IHC practice). The HPA finding applies to these cell types, not every cell in their tissues. |
| A pale, even deposit covers cells and extracellular areas without a clear compartment pattern. | Treat the diffuse deposit as background until controls establish a specific signal (general IHC practice). Review blocking, washing and detection conditions, then ask whether cytoplasmic or membrane staining remains in an expected cell population (HPA: tissue IHC; UniProt Q5GJ75 localization). Do not score the uniform haze as positive. |
| No staining is visible in glandular cells of a planned positive section. | Colon, duodenum and endometrial glandular cells are scored High in HPA tissue IHC, making an absent signal a reason to check the run (HPA: tissue IHC). Confirm the relevant cells are present, then inspect primary-antibody and chromogenic detection steps (general IHC practice). A failed run cannot establish true absence. |
💡Expected TNFAIP8L3 appearanceCall a section positive when identifiable smooth muscle or HPA High glandular cells show cell-associated cytoplasmic staining, with possible membrane accentuation; isolated nuclear-only signal or diffuse chromogen without that pattern warrants review (HPA: tissue IHC; UniProt Q5GJ75 localization).
| Tissue and cell choice | Use cell-level expectations: smooth muscle is described as most abundant, whereas kidney tubule cells and ovarian follicle cells are scored High; adipocytes are Not detected (HPA: tissue IHC). A whole tissue label alone cannot identify which cells should stain. |
| Localization and topology | UniProt lists cytoplasm and cell membrane, with movement toward the plasma membrane on PDGF activation, but annotates no transmembrane segment (UniProt Q5GJ75). Membrane accentuation can fit the record; an activation-dependent shift should not be presumed from an unstated specimen history. |
| Antibody evidence | HPA077484 is listed as IHC Approved; the tissue IHC profile is also Approved, with external verification pending (HPA: antibody validation; HPA: tissue IHC reliability). These labels support using the displayed pattern as a comparator, while leaving unexpected staining to be checked with controls. |
| IF/ICC question: should nuclear signal be expected? | HPA reports cytosol as the supported main ICC-IF location and nucleoplasm as an approved additional location (HPA: subcellular ICC-IF). That ICC-IF observation does not make nuclear-only staining the expected paraffin IHC result; assess each assay against its own reported pattern. |
| Fixation and retrieval evidence | Target-specific fixation sensitivity and retrieval requirements are unreported in the supplied UniProt and HPA records. Document the conditions used and interpret controls within that run (general IHC practice); do not infer a TNFAIP8L3 fixation effect from tissue staining levels. |
| Situation | Likely cause | Next action |
|---|---|---|
| The planned positive section is blank. | The expected cell population may be absent from the examined area, or the IHC run may have failed (HPA: High glandular populations; general IHC practice). | Locate the relevant cells, check a positive control, and review primary-antibody incubation and chromogenic detection before scoring absence (general IHC practice). |
| Adipocytes stain as strongly as the intended positive cells. | This conflicts with the Not detected adipocyte observation; cross-reactivity or detection background is possible (HPA: adipose tissue; general IHC practice). | Inspect a no-primary control and compare cell-level staining in the same run before accepting adipocyte signal (general IHC practice). |
| The entire section has diffuse brown staining. | A compartment-free deposit is consistent with nonspecific background or endogenous chromogenic activity (general IHC practice). | Check the no-primary control, blocking and wash steps; score only interpretable cell-associated staining after background is resolved (general IHC practice). |
| Signal appears exclusively nuclear in IHC. | The result differs from the reported cytoplasmic and membrane tissue pattern (HPA: tissue IHC; UniProt Q5GJ75 localization). | Review controls and detection, then reassess the cellular compartment. Keep the additional nucleoplasmic ICC-IF finding separate from the paraffin IHC call (HPA: subcellular ICC-IF). |
| A low-staining population looks nearly blank. | Lung macrophages and stomach glandular cells are scored Low, so their faint signal may be difficult to distinguish from background (HPA: tissue IHC). | Check the run using a High-scored cell population, then evaluate the low signal against local background and controls (HPA: tissue IHC; general IHC practice). |
| Membrane staining varies between positive cells. | UniProt reports cytoplasmic and membrane localization and PDGF-associated movement to the plasma membrane; the specimen's activation state is unspecified (UniProt Q5GJ75). | Record cytoplasmic and membrane staining separately. Do not assign the variation to PDGF activation without independent specimen information (UniProt Q5GJ75; general IHC practice). |
Experiment evidence
IHC & IF images showing TNFAIP8L3 staining
PA2257 · IHCAnti-TNFAIP8L3 antibody, PA2257, IHC(P) IHC(P): Human Intestinal Cancer Tissue
PA2257 · IHCAnti-TNFAIP8L3 antibody, PA2257, IHC(P) IHC(P): Human Mammary Cancer Tissue
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000183578-TNFAIP8L3/tissue/cerebral+cortex#imid_20683496
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000183578-TNFAIP8L3/tissue/colon#imid_20683414
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000183578-TNFAIP8L3/tissue/liver#imid_20683431
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000183578-TNFAIP8L3/tissue/kidney#imid_20683432
HPA · ICC-IF HUVEC/TERT2Human Protein Atlas immunofluorescence, HUVEC/TERT2, antibody HPA077484; green = TNFAIP8L3, blue = nucleus, red = microtubules.
HPA · ICC-IF HUVEC/TERT2Human Protein Atlas immunofluorescence, HUVEC/TERT2, antibody HPA077484; green = TNFAIP8L3, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for TNFAIP8L3 IHC & IF
🧪Run colon first and score its glandular cells, where TNFAIP8L3 staining is High (HPA: Colon, glandular cells). Use cervix glandular cells as the negative tissue (Not detected; HPA: Cervix, glandular cells); on the colon slide, assess surrounding non-glandular cells for background staining, but do not assume they are target-negative because their status is unreported in the supplied HPA row.
Positive control tissue: Bronchus (Ciliated cells (ciliary rootlets), HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show TNFAIP8L3 in HUVEC/TERT2, HeLa, U-251MG, with annotated localisation: Cytosol (supported) (HPA subcellular).
Technical controls: Include a no-primary (secondary-only) slide, an isotype control matched to the primary antibody’s host species and clonality, and a knockout specimen if available; peptide blocking can provide an additional specificity check. For chromogenic colon IHC, quench endogenous peroxidase and assess residual signal with the no-primary slide; block endogenous biotin if using avidin–biotin detection (standard IHC practice).
⚠️Feasibility: A target-specific fixation window and its effect are unreported, and the selected PA2257 paraffin-section caption does not state a fixative (PA2257 tissue-IHC caption). Target-specific dependence on antigen retrieval is also unreported; optimize retrieval using standard paraffin IHC controls, and do not assume frozen sections or IF are easier from the supplied evidence (PA2257 tissue-IHC caption; HPA: ICC-IF subcellular images). In colon, mucus or endogenous peroxidase can complicate interpretation of chromogenic staining (standard IHC practice).
HPA tissue IHC evidence for TNFAIP8L3
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Antibody staining in cells/structures not annotated, view images. 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 |
|---|---|---|---|---|
| Bronchus | Ciliated cells (ciliary rootlets) | High | Protein (IHC) | HPA → |
| Colon | Glandular cells | High | Protein (IHC) | HPA → |
| Duodenum | Glandular cells | High | Protein (IHC) | HPA → |
| Endometrium | Glandular cells | High | Protein (IHC) | HPA → |
| Epididymis | Glandular cells | High | 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 → |
| Cerebral cortex | Endothelial cells | Not detected | Protein (IHC) | HPA → |
| Cervix | Glandular cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced TNFAIP8L3 IHC Tips
Troubleshoot TNFAIP8L3 staining by comparing signal with expected cytoplasmic and membrane patterns, appropriate tissue controls, and matched processing conditions (UniProt Q5GJ75 localisation; HPA tissue IHC).
How should I adjust retrieval when TNFAIP8L3 staining is weak in paraffin sections?
Start with citrate buffer at pH 6.0, heated to 95–98 °C for 20 min (page retrieval rule: cytoplasmic / membrane antigen). Keep section thickness, cooling time and antibody incubation consistent while comparing a weak section with a positive control (standard IHC practice). Colon glandular cells provide a high-staining reference, although their staining does not establish optimal retrieval for every specimen (HPA: High in colon glandular cells). If signal remains weak, test a different retrieval buffer or duration on adjacent sections as a fallback, recording tissue damage and background (standard IHC practice). Assess cytoplasmic and membrane staining separately because both locations are reported for TNFAIP8L3 (UniProt Q5GJ75 localisation).
Could fixation explain weak or uneven TNFAIP8L3 staining?
Target-specific fixation sensitivity for TNFAIP8L3 is unknown from the supplied evidence; the selected PA2257 caption identifies paraffin tissue IHC but does not state its fixative (PA2257 tissue-IHC caption). Compare sections with documented fixation and processing histories before attributing a weak result to antigen loss (standard IHC practice). Use the same retrieval, primary antibody incubation and detection conditions across that comparison so fixation is the main changing variable (standard IHC practice). Look for staining gradients between the tissue edge and center, and inspect morphology for processing damage (standard IHC practice). Do not infer fixation tolerance from the reported tissue pattern or protein features (HPA tissue IHC; UniProt Q5GJ75 record).
Which staining compartments should count as plausible TNFAIP8L3 signal?
Score cytoplasmic and cell membrane staining as plausible tissue IHC patterns, since both are reported for TNFAIP8L3 (UniProt Q5GJ75 localisation; HPA tissue IHC). Cytosolic signal is also supported in cell imaging, while nucleoplasmic localisation is an additional approved observation rather than the dominant reported location (HPA subcellular). Record nuclear, cytoplasmic and membrane staining separately instead of pooling every stained pixel into one score (standard IHC practice). A membrane-enriched pattern may vary with signaling state because PDGF activation can move TNFAIP8L3 from cytoplasm to plasma membrane (UniProt Q5GJ75 localisation). Compare the same cell type and processing conditions before treating compartment shifts as a biological difference (standard IHC practice).
How do I investigate an unexpected staining pattern if the antibody epitope is unclear?
Check the catalog antibody’s stated immunogen and epitope region before interpreting a compartment-specific result; neither is specified in the supplied evidence (catalog antibody: epitope information not supplied). The provided record lists 0 isoforms, no annotated domains and no transmembrane segment, so those annotations do not explain an unexpected pattern (UniProt Q5GJ75 record). It also lists no signal peptide, propeptide, glycosylation sites or modified residues; these annotations do not establish how fixation exposes the epitope (UniProt Q5GJ75 record). Compare adjacent sections with and without primary antibody and review the expected cell pattern (standard IHC practice). Treat an epitope-dependent explanation as provisional until antibody-specific evidence supports it (standard IHC practice).
How can I check TNFAIP8L3 localisation by multiplex IF alongside tissue IHC?
Pair TNFAIP8L3 with a validated marker for the expected cell type, such as a smooth muscle marker when examining smooth muscle, and verify each channel alone (HPA tissue IHC: strongest abundance in smooth muscle; standard IF practice). Choose fluorophores after measuring tissue autofluorescence in unstained sections, and favor a well-separated channel for weak TNFAIP8L3 signal (standard IF practice). Decide permeabilisation from the mapped epitope: a cytosolic epitope needs access through the membrane, whereas a proven extracellular epitope may be tested without detergent (standard IF practice). No transmembrane segment or epitope orientation is supplied, so do not assume surface accessibility (UniProt Q5GJ75 topology; catalog antibody: epitope information not supplied). Check whether IF cytosolic staining agrees with the supported cell-imaging location (HPA subcellular: cytosol supported).
What should I check when chromogenic TNFAIP8L3 staining is diffuse or granular?
Inspect a section processed without primary antibody to identify detection-system and tissue background before changing antibody concentration (standard IHC practice). Block endogenous peroxidase for a peroxidase-based assay, then compare DAB development times on matched sections (standard chromogenic IHC practice). Examine pigment, folds, damaged edges and necrotic areas at the same exposure and counterstain settings used for scored regions (standard IHC practice). Diffuse staining in every cell type deserves scrutiny because HPA reports variable tissue and cell staining, including no detection in adipocytes (HPA tissue IHC: adipocytes not detected). Confirm that retained signal has plausible cytoplasmic or membrane distribution before calling it TNFAIP8L3 (UniProt Q5GJ75 localisation).
How should I quantify TNFAIP8L3 staining across heterogeneous paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring because TNFAIP8L3 is reported in cytoplasm and at the cell membrane (UniProt Q5GJ75 localisation; HPA tissue IHC). For chromogenic IHC, report an H-score from staining intensity and the percentage of positive cells, with the scoring thresholds fixed across slides (standard IHC practice). If measuring spatial signal, report positive-cell density per mm² of viable, annotated tissue and state how excluded regions were handled (standard IHC practice). Normalize comparisons to the same cell type, viable area, batch controls and detection settings rather than raw whole-slide brown area (standard IHC practice). Report cytoplasmic and membrane results separately when a redistribution question is being tested (UniProt Q5GJ75 localisation; standard IHC practice).
How can I distinguish true TNFAIP8L3 staining from section artefacts?
Favor reproducible cytoplasmic or membrane staining in intact cells, consistent with the reported tissue distribution and protein localisation (HPA tissue IHC; UniProt Q5GJ75 localisation). Compare a high-staining reference, such as colon glandular cells, with a low or undetected reference assessed under the same conditions (HPA: High in colon glandular cells; HPA: adipocytes not detected; standard IHC practice). Investigate staining confined to section edges, folds or necrosis as a processing artefact, and use a no-primary control to assess endogenous enzyme or detection background (standard IHC practice). Strong staining in an unexpected cell or solely nuclear compartment warrants separate verification before biological interpretation (HPA subcellular: cytosol supported, nucleoplasm additional; standard IHC practice). A membrane shift is biologically plausible under PDGF activation, but requires matched experimental evidence (UniProt Q5GJ75 localisation).
Boster reagents
Best TNFAIP8L3 / Tumor necrosis factor alpha-induced protein 8-like protein 3 IHC Antibodies
PA2257 is a human-reactive IHC antibody with paraffin-section images of human intestinal and mammary cancer tissue (catalog: PA2257 reactivity and IHC image captions).
Real IHC data
Anti-TNFAIP8L3 Antibody ®
Cat # PA2257
PA2257 is listed for human IHC (catalog: PA2257 applications/reactivity). Its IHC(P) captions show human intestinal cancer tissue and human mammary cancer tissue (catalog: PA2257 IHC image captions).
Which to pick: Choose PA2257 for paraffin-section tissue IHC; its listed concentration is 0.5–1 μg/ml (catalog: PA2257 IHC image captions; datasheet: 0.5-1μg/ml). It has a rabbit host, but clonality and the tissue fixative are unreported (catalog: PA2257 host/clone and IHC image captions). No IF/ICC or cross-species pick is supported by this catalog: PA2257 lists human reactivity and IHC/WB applications, with no IF images (catalog: PA2257 reactivity/applications/if_image_alts).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q5GJ75 (TP8L3_HUMAN, Tumor necrosis factor alpha-induced protein 8-like protein 3).
- Human Protein Atlas. TNFAIP8L3 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. TNFAIP8L3 subcellular location (ICC-IF): Mainly localized to the cytosol. In addition localized to the nucleoplasm..
- Human Protein Atlas. TNFAIP8L3 antibody validation summary (1 antibodies).
- The Correlation and Clinicopathological Significance of TNFAIP8L3 and RAC1 Expression in Lung Adenocarcinoma. Genetics research 2025 — PMC12404833.
- TIPE3 promotes breast cancer progression and metastasis via the AKT-GSK3β-β-catenin/Snail pathway. Translational cancer research 2025 — PMC12605601.
- TIPE3 differentially modulates proliferation and migration of human non-small-cell lung cancer cells via distinct subcellular location. BMC cancer 2018 — PMC5840720.
- TIPE3 promotes non-small cell lung cancer progression via the protein kinase B/extracellular signal-regulated kinase 1/2-glycogen synthase kinase 3β-β-catenin/Snail axis. Translational lung cancer research 2021 — PMC7947417.
- PubMed PMID:14702039 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
- PubMed PMID:25479791 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for TNFAIP8L3 (Tumor necrosis factor alpha-induced protein 8-like protein 3). Expected localisation Cytoplasmic and membranous tissue staining (HPA tissue IHC). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: PA2257 at 0.5-1μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Bronchus (Ciliated cells (ciliary rootlets)); negative Adipose tissue. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Tissue sections; selected-image fixative not specified (standard IHC workflow); 2. Fixation: Image fixative and duration unreported (datasheet PA2257); 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-TNFAIP8L3, 0.5-1μg/ml (datasheet PA2257); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across paraffin sections. (standard IHC practice; not target-specific); Caveat: PDGF activation can shift cytoplasmic signal to the membrane (UniProt); Isoform / epitope: No isoforms or processing annotated; epitope region unreported (UniProt); Regulation: Expression regulation unreported (UniProt). Evidence: UniProt Q5GJ75, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the cytosol. In addition localized to the…, Datasheet PA2257, PMC12404833, PMC5840720. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/TNFAIP8L3/immunohistochemistry-ihc-guide.