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TMOD3 / Tropomodulin-3 · IHC design guide
Design Immunohistochemistry for TMOD3
Plan chromogenic IHC on paraffin sections using the IHC-validated antibody at 1:100–1:200 (datasheet: A07431). Assess cytoplasmic staining while treating tissue-level results as provisional because antibody staining and RNA show low consistency (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
TMOD3 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); cytoskeleton (UniProt) | |
| Staining pattern | Cytoplasmic staining in most tissues, including glandular cells (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Adrenal gland+4 more · see all | |
| Negative control | Skeletal muscle |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation conditions consistent (standard IHC practice; not target-specific) | |
| Caveat | Antibody staining and RNA show low consistency (HPA tissue IHC) | |
| Regulation | Specific expression regulation is unreported (UniProt) | |
| Isoform / epitope | No annotated isoforms; one cytoplasmic chain, not extracellular (UniProt) |
Section 1
Recommended TMOD3 IHC & IF Protocols
The catalog antibody protocol is accompanied by one published TMOD3 IHC protocol using pancreatic adenocarcinoma tissue microarrays (PMC11165712 methods).
| Sample | Paraffin-embedded human esophagus tissue; fixative not specified (datasheet A07431) |
| Fixation | Image fixative and duration unreported (datasheet A07431); 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-TMOD3, 1:100-1:200 (datasheet A07431) |
| 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 | TMOD3-positive staining in glandular cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in most 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); the published pancreatic protocol does not specify retrieval (PMC11165712 methods).
Section 2
What Is the Expected TMOD3 Staining Pattern?
TMOD3 is a cytoplasmic, actin-associated protein with no transmembrane segment (UniProt Q9NYL9: localization, function, topology). Expect cytoplasmic staining in many tissue cell types, including glandular, hematopoietic, respiratory epithelial, glial and neuronal cells (HPA tissue IHC: cytoplasmic expression in most tissues; listed cells High). Interpret these patterns cautiously: HPA rates the tissue IHC antibody Approved but reports low staining–RNA consistency and pending external verification (HPA tissue IHC: reliability).
| Cytoplasmic staining in adrenal or appendix glandular cells, or marrow hematopoietic cells (HPA tissue IHC: High). | This matches the reported compartment and high-staining cell types (HPA tissue IHC: cytoplasmic; listed cells High). Compare the cells on the slide with the tissue annotation; an Approved rating does not resolve the reported staining–RNA inconsistency (HPA tissue IHC: reliability). |
| Predominantly nuclear or membrane-only staining, without a convincing cytoplasmic pattern (UniProt Q9NYL9: localization, topology). | This conflicts with cytoplasmic IHC and cytoskeletal localization (HPA tissue IHC: cytoplasmic; UniProt Q9NYL9: cytoskeleton). Treat it as suspect and check morphology, staining controls and detection artifacts before assigning a new TMOD3 location (general IHC interpretation practice). |
| Strong staining confined to unexpected cells while the reported positive cells remain unstained (HPA tissue IHC: listed High cells). | Possible cross-reactivity or endogenous chromogenic activity needs investigation (general IHC interpretation practice). Broad expression alone cannot verify an unexpected cell pattern, especially with low staining–RNA consistency (UniProt Q9NYL9: ubiquitous; HPA tissue IHC: reliability). |
| Color covers extracellular spaces, section edges or most cells without clear cytoplasmic boundaries (general IHC interpretation practice). | Diffuse background limits compartment and cell-type scoring (general IHC interpretation practice). Compare a primary-antibody omission control, then assess blocking, washing and chromogen development; background alone is not evidence of TMOD3 expression (general IHC practice). |
| No cytoplasmic signal in a reported High cell population, such as breast glandular cells (HPA tissue IHC: breast glandular cells High). | The run may have insufficient usable signal, or this specimen may differ from the HPA example (general IHC interpretation practice). Check section quality and assay controls before calling it negative; HPA reports variable agreement between staining and RNA (HPA tissue IHC: reliability). |
💡Expected TMOD3 appearanceA convincing positive shows cell-resolved cytoplasmic chromogen in reported High populations, such as appendix glandular cells (HPA tissue IHC: cytoplasmic; appendix High); isolated nuclear, membrane-only or structureless diffuse color is suspect (UniProt Q9NYL9: localization, topology; general IHC interpretation practice).
| Compartment and protein role (UniProt Q9NYL9: localization, function) | TMOD3 is cytoplasmic and binds actin and tropomyosin (UniProt Q9NYL9: subunit). Score staining within cells rather than requiring a membrane rim; UniProt reports no transmembrane segment (UniProt Q9NYL9: topology). |
| Tissue distribution (HPA tissue IHC: profile and cell-level staining) | Cytoplasmic expression occurs in most tissues, yet levels differ by cell type (HPA tissue IHC: profile). Skeletal myocytes are Not detected, while cardiomyocytes and smooth muscle cells are Low (HPA tissue IHC: negative and low). |
| Confidence of the tissue pattern (HPA tissue IHC: reliability; HPA antibodies: HPA001849) | The listed antibody has Approved IHC status, while the tissue profile notes low staining–RNA consistency and pending external verification (HPA antibodies: IHC Approved; HPA tissue IHC: reliability). Interpret unexpected results with controls. |
| IF/ICC localization (HPA subcellular ICC-IF: approved locations) | The separate IF/ICC evidence places TMOD3 mainly in cytosol and additionally on actin filaments (HPA subcellular ICC-IF: approved). It supports a localization comparison, without establishing an IHC staining intensity or retrieval setting. |
| Situation | Likely cause | Next action |
|---|---|---|
| Positive tissue has weak or absent cytoplasmic color (HPA tissue IHC: reported High cells). | The IHC run may have inadequate signal, or the specimen may differ from the reference pattern (general IHC practice; HPA tissue IHC: reliability). | Check section integrity and controls, then review the assay's validated retrieval, antibody dilution and detection conditions (general IHC practice). No TMOD3-specific retrieval effect is supplied. |
| Nuclear or membrane-only color dominates (UniProt Q9NYL9: cytoplasmic, no transmembrane segment). | The compartment disagrees with the documented localization; nonspecific or detection staining is possible (HPA tissue IHC: cytoplasmic; general IHC practice). | Inspect cellular boundaries and compare a primary-antibody omission control before scoring the color as TMOD3 (general IHC interpretation practice). |
| Diffuse color obscures individual cells (general IHC interpretation practice). | Background from the staining workflow or endogenous chromogenic activity may obscure true signal (general IHC practice). | Compare omission and detection controls; review blocking, washes and chromogen development using the assay's established workflow (general IHC practice). |
| Skeletal myocytes stain as strongly as a reported High population (HPA tissue IHC: skeletal myocytes Not detected). | The result conflicts with HPA's tissue observation, although that observation has limited consistency with RNA data (HPA tissue IHC: negative; reliability). | Confirm cell identity and review controls and specimen context before treating this contrast as specificity evidence (general IHC interpretation practice). |
| A cell population expected to be Low appears faint (HPA tissue IHC: cardiomyocytes, smooth muscle cells Low). | Faint staining can agree with the reported level; visual intensity varies with assay conditions (HPA tissue IHC: Low; general IHC practice). | Score against an appropriate positive control from the same run and preserve the cell-type distinction (general IHC interpretation practice). |
| What localization should a separate IF/ICC assay show? | HPA reports cytosol as the main location and actin filaments as an additional location (HPA subcellular ICC-IF: approved). | Use that pattern when interpreting the separate IF/ICC assay; this IHC section supplies no IF/ICC protocol conditions (HPA subcellular ICC-IF: localization only). |
Experiment evidence
IHC & IF images showing TMOD3 staining
A07431 · IHCImmunohistochemistry of paraffin-embedded human esophagus using TMOD3 antibody at dilution of 1:100 .Perform microwave antigen retrieval wit
A07431 · IHCImmunohistochemistry of paraffin-embedded mouse brain using TMOD3 antibody at dilution of 1:100 .Perform microwave antigen retrieval with 10
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000138594-TMOD3/tissue/cerebral+cortex#imid_917336
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000138594-TMOD3/tissue/colon#imid_917262
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000138594-TMOD3/tissue/liver#imid_917271
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000138594-TMOD3/tissue/kidney#imid_917272
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA001849; green = TMOD3, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA001849; green = TMOD3, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for TMOD3 IHC & IF
🧪Run bronchus first: respiratory epithelial cells should stain (HPA: High in respiratory epithelial cells). Use skeletal muscle as the biological tissue comparator (HPA: Not detected in myocytes); any unstained cells within the bronchus section can help assess background, but should not be assumed to be true TMOD3-negative cells (UniProt Q9NYL9: ubiquitous tissue specificity).
Positive control tissue: Adrenal gland (Glandular cells, HPA High)
Negative control tissue: Skeletal muscle (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show TMOD3 in A-431, U-251MG, U2OS, with annotated localisation: Cytosol (approved) (HPA subcellular).
Technical controls: Include no-primary (secondary-only) and host-species- and clonality-matched isotype controls, plus a TMOD3 knockout sample or a validated peptide-block control (standard IHC practice). Quench endogenous peroxidase for chromogenic detection; for IF, inspect an unstained bronchus section for autofluorescence (standard IHC/IF practice).
⚠️Feasibility: A target-specific fixation window and fixation effect are unreported in the supplied evidence; the selected A07431 paraffin-section caption does not state a fixative (A07431 tissue-IHC caption). That caption uses microwave retrieval in 10 mM PBS, pH 7.2, at 1:100 for human esophagus, but does not establish whether TMOD3 staining depends on retrieval in bronchus (A07431 tissue-IHC caption). The supplied evidence does not establish that frozen sections or IF are easier; bronchial mucus can retain staining reagents and should be assessed against the no-primary control (standard IHC practice).
HPA tissue IHC evidence for TMOD3
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Low consistency between antibody staining and RNA expression data. 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 | High | Protein (IHC) | HPA → |
| Appendix | Glandular cells | High | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | High | Protein (IHC) | HPA → |
| Breast | Glandular cells | High | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Skeletal muscle | Myocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced TMOD3 IHC Tips
Troubleshoot TMOD3 chromogenic IHC in paraffin sections by checking retrieval, cytoplasmic localisation, controls and cell-specific scoring.
Which retrieval conditions should I try first when TMOD3 staining is weak?
Start with citrate buffer at pH 6.0, heated to 95–98 °C for 20 minutes (page retrieval rule). If staining remains weak, compare the selected antibody’s documented method: microwave retrieval in 10 mM PBS, pH 7.2, on paraffin-embedded human esophagus at 1:100 (A07431 tissue-IHC caption). Keep section thickness, detection and exposure to chromogen consistent while comparing retrieval conditions (standard IHC practice). Check whether improved signal appears in cytoplasm rather than as widespread nuclear staining, because TMOD3 is a cytoskeletal protein and HPA reports mainly cytosolic localisation (UniProt Q9NYL9; HPA subcellular).
How should I troubleshoot fixation-related loss of TMOD3 signal?
The supplied evidence does not establish TMOD3-specific sensitivity to any fixative or fixation duration; the selected paraffin-section caption does not state a fixative (A07431 tissue-IHC caption). Record the actual fixative, duration and tissue processing history before comparing sections, since these variables can affect antigen accessibility in IHC (standard IHC practice). Compare matched sections with the same citrate retrieval at pH 6.0, 95–98 °C for 20 minutes and identical detection conditions (page retrieval rule; standard IHC practice). If signal differs, describe an association with processing conditions rather than assigning a TMOD3-specific fixation mechanism without a controlled comparison (standard IHC practice).
What staining pattern is plausible for TMOD3 in tissue sections?
Expect predominantly cytoplasmic staining: TMOD3 is annotated in the cytoplasm and cytoskeleton, while HPA describes cytoplasmic expression in most tissues (UniProt Q9NYL9; HPA tissue IHC). Cytosolic signal with some filament-associated detail is plausible because HPA reports approved cytosolic and additional actin-filament localisation (HPA subcellular). TMOD3 has no annotated transmembrane segment, so isolated crisp membrane staining needs independent validation before assignment to TMOD3 (UniProt Q9NYL9 topology; standard IHC practice). Compare cells within the same section using a consistent counterstain and inspect morphology at high power; do not treat staining intensity alone as proof of subcellular localisation (standard IHC practice).
Can isoforms or epitope accessibility explain discrepant TMOD3 IHC results?
The supplied record lists 0 annotated isoforms, one 352-residue chain and no annotated domains, so it provides no basis for assigning divergent IHC patterns to a named isoform (UniProt Q9NYL9). It lists phosphoserine at position 25, but no antibody epitope or phosphorylation sensitivity is supplied (UniProt Q9NYL9; supplied antibody evidence). Compare adjacent sections under the same retrieval and antibody conditions before attributing a difference to epitope accessibility (standard IHC practice). If possible, validate the pattern with an independently characterised antibody or a suitable loss-of-target control; record which epitopes those reagents recognise before interpreting disagreement (standard IHC practice).
How can IF help assess an ambiguous TMOD3 IHC pattern?
Use IF as an orthogonal localisation check and multiplex TMOD3 with a validated marker for the cell type being assessed, so overlapping cells can be identified (standard IF practice). HPA reports TMOD3 mainly in cytosol, with additional actin-filament localisation, providing a pattern to compare with chromogenic IHC (HPA subcellular). Choose a fluorophore channel with low tissue autofluorescence and include single-stain and unstained controls before interpreting overlap (standard IF practice). Because TMOD3 has no transmembrane segment and is cytoplasmic, permeabilise cells for access to the intracellular epitope; optimise detergent conditions against preservation of cell structure (UniProt Q9NYL9 topology and localisation; standard IF practice).
How do I reduce diffuse background without losing cytoplasmic TMOD3 signal?
First distinguish expected cytoplasmic signal from staining that also appears in cell-free areas or on a reagent-only control (HPA tissue IHC; standard IHC practice). Use an appropriate protein block, thorough washes and a no-primary control to assess nonspecific detection (standard IHC practice). For peroxidase-based chromogenic IHC, block endogenous peroxidase before DAB development and compare development times across matched sections (standard IHC practice). The selected antibody was shown on paraffin-embedded human esophagus at 1:100; treat that dilution as a documented starting point for A07431, then titrate if background obscures cytoplasmic detail (A07431 tissue-IHC caption; standard IHC practice).
How should I score TMOD3 staining across heterogeneous tissue? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and analysis region before scoring, then report cytoplasmic intensity and the percentage of positive cells; an H-score combines those two observations (standard IHC practice; HPA tissue IHC). For spatial comparisons, report positive-cell density per mm² of viable tissue and state the area excluded for folds or damage (standard IHC practice). Normalise counts to the relevant cell population or analysed viable area, rather than to the entire image when tissue composition differs (standard IHC practice). Keep retrieval, imaging and scoring thresholds fixed across cases, and interpret modest differences cautiously because HPA flags low consistency between antibody staining and RNA expression (standard IHC practice; HPA tissue IHC reliability).
When should an apparent TMOD3-positive IHC result be questioned?
A credible result should fit predominantly cytoplasmic localisation in intact cells; HPA also reports cytosolic and actin-filament localisation (UniProt Q9NYL9; HPA subcellular). Question isolated nuclear or sharp membrane staining, especially when the no-primary control is similar, because neither pattern matches the supplied localisation evidence (UniProt Q9NYL9; HPA subcellular; standard IHC practice). Check whether signal tracks section edges, necrotic areas or endogenous enzyme activity before calling cells positive (standard IHC practice). Cell identity matters: HPA reports high staining in bone-marrow hematopoietic cells but no detection in skeletal-muscle myocytes, while its tissue-IHC profile carries a low-consistency warning (HPA tissue IHC).
Boster reagents
Best TMOD3 / Tropomodulin-3 IHC Antibodies
A07431 has IHC images from paraffin-embedded human esophagus and mouse brain (A07431 image captions); Rat reactivity is listed without a tissue image (A07431 catalog).
Real IHC data
Anti-Tropomodulin-3 TMOD3 Antibody
Cat # A07431
A07431 will render with IHC images of paraffin-embedded human esophagus and mouse brain, each stained at 1:100 after microwave retrieval in 10 mM PBS, pH 7.2 (A07431 image captions). Its catalog lists IHC and Human, Mouse and Rat reactivity; no IF/ICC image or dilution is supplied (A07431 catalog).
Which to pick: Choose A07431 for paraffin-section tissue IHC: it is a rabbit antibody with human esophagus and mouse brain image evidence and a listed IHC dilution of 1:100–1:200 (A07431 catalog; A07431 image captions). For cross-species work, Human, Mouse and Rat reactivity is listed, while the IHC images document Human and Mouse tissues only (A07431 catalog; A07431 image captions). No IF/ICC-validated SKU is provided, and the paraffin-section captions do not report a fixative (A07431 catalog; A07431 image captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q9NYL9 (TMOD3_HUMAN, Tropomodulin-3).
- Human Protein Atlas. TMOD3 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. TMOD3 subcellular location (ICC-IF): Mainly localized to the cytosol. In addition localized to the actin filaments..
- Human Protein Atlas. TMOD3 antibody validation summary (1 antibodies).
- Clinicopathological Characteristics, Prognosis, and Correlated Tumor Cell Function of Tropomodulin-3 in Pancreatic Adenocarcinoma. Combinatorial chemistry & high throughput screening 2024 — PMC11165712.
- YWHAG promotes bladder cancer metastasis by regulating TMOD3 to activate ERK1/2 and JNK phosphorylation in the MAPK pathway. Journal of translational medicine 2024 — PMC11687020.
- PubMed PMID:10662549 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for TMOD3 (Tropomodulin-3). Expected localisation Cytoplasmic tissue staining (HPA tissue IHC); cytoskeleton (UniProt). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: A07431 at 1:100-1:200 (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Glandular cells); negative Skeletal muscle. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Paraffin-embedded human esophagus tissue; fixative not specified (datasheet A07431); 2. Fixation: Image fixative and duration unreported (datasheet A07431); 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-TMOD3, 1:100-1:200 (datasheet A07431); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation conditions consistent (standard IHC practice; not target-specific); Caveat: Antibody staining and RNA show low consistency (HPA tissue IHC); Isoform / epitope: No annotated isoforms; one cytoplasmic chain, not extracellular (UniProt); Regulation: Specific expression regulation is unreported (UniProt). Evidence: UniProt Q9NYL9, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the cytosol. In addition localized to the actin…, Datasheet A07431, PMC11165712. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/TMOD3/immunohistochemistry-ihc-guide.