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ID1 / DNA-binding protein inhibitor ID-1 · IHC design guide
Design Immunohistochemistry for ID1
Plan ID1 paraffin IHC around cytoplasmic and nuclear staining across tissues (HPA tissue IHC). Bronchial respiratory epithelium and pancreatic exocrine cells show high staining and can serve as positive controls (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
ID1 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 nuclear tissue staining (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic and nuclear staining across tissues (HPA tissue IHC) | |
| Antigen retrieval | Tris-EDTA pH 9.0 HIER, 95–98 °C, 20 min (rule: nuclear antigen) | |
| Positive control | Bronchus+4 more · see all | |
| Negative control | Adipose tissue+1 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation conditions consistent across sections. (standard IHC practice; not target-specific) | |
| Caveat | Adipocytes may be unstained despite broad tissue expression (HPA tissue IHC) | |
| Regulation | No expression stimulus specified (UniProt) | |
| Isoform / epitope | 2 isoforms (ID-A/ID-B); epitope coverage unknown (UniProt) |
Section 1
Recommended ID1 IHC & IF Protocols
The catalog antibody protocol is followed by published ID1 IHC methods for gastric, bladder, and lung cancer samples (PMC2714249; PMC4927902; PMC3567999).
| Sample | Paraffin-embedded human kidney tissue; fixative not specified (datasheet M00945) |
| Fixation | Image fixative and duration unreported (datasheet M00945); 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 Tris-EDTA buffer, pH 9.0, 20 min at 95–98 °C (standard rule: nuclear 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 monoclonal (clone ACC-9) anti-ID1, 1:50 (datasheet M00945) |
| 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 | ID1-positive staining in respiratory epithelial cells of bronchus (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic and nuclear expression in all tissues. No signal in the no-primary control. |
💡Decision noteStart with the catalog antibody’s Tris-EDTA pH 9.0 retrieval at 95–98 °C for 20 min (page antigen retrieval); adjust for the published antibody and sample.
Section 2
What Is the Expected ID1 Staining Pattern?
ID1 localizes to the nucleus and cytoplasm (UniProt P41134; HPA: Approved tissue IHC). Expect high staining in bronchial respiratory epithelial cells, pancreatic exocrine cells, placental trophoblasts, chondrocytes and tonsillar non-germinal center cells (HPA: High). A membrane-only pattern is unexpected because ID1 has no transmembrane segment (UniProt P41134 topology).
| Nuclear and cytoplasmic signal in bronchial respiratory epithelial cells (HPA: High). | Fits the reported tissue pattern (HPA: Approved tissue IHC). |
| Membrane-only staining. | Suspect artefact; ID1 is intracellular and lacks a transmembrane segment (UniProt P41134). |
| Signal confined to adipocytes. | Check cross-reactivity or endogenous detection activity; adipocytes are Not detected (HPA). |
| Diffuse haze across cells and blank areas. | Assess nonspecific background with blocking, wash and detection controls (general IHC practice). |
| No signal in bronchial respiratory epithelial cells (HPA: High). | Review the assay and positive control; this cell type stains High in HPA tissue IHC (HPA). |
💡Expected ID1 appearanceCall the result positive when nuclear and cytoplasmic staining is high in bronchial respiratory epithelial or pancreatic exocrine cells (HPA: High); membrane-only staining is suspect (UniProt P41134 topology).
| Control tissue | Bronchial epithelium is High; adipocytes are Not detected (HPA tissue IHC). |
| IHC antibody validation | CAB025915 is Approved for IHC; the other listed antibodies have no IHC rating (HPA antibodies). |
| Isoforms and processing | ID-A and ID-B are listed; no signal peptide or propeptide is listed (UniProt P41134). |
| IF/ICC expectation? | Nucleoplasmic signal is reported (HPA: enhanced ICC-IF); consult the separate IF/ICC guide. |
| Situation | Likely cause | Next action |
|---|---|---|
| No signal in bronchial respiratory epithelium (HPA: High). | Possible retrieval or detection failure (general IHC practice). | Check an on-slide positive control and the retrieval and detection steps (general IHC practice). |
| Membrane-only staining. | Possible nonspecific signal; ID1 lacks a transmembrane segment (UniProt P41134). | Review antibody specificity and a no-primary control (general IHC practice). |
| Adipocytes stain strongly (HPA: Not detected). | Possible cross-reactivity or endogenous detection activity (general IHC practice). | Compare no-primary and detection controls (general IHC practice). |
| Diffuse haze across the section. | Possible nonspecific antibody binding (general IHC practice). | Review blocking, antibody dilution and washes (general IHC practice). |
| Signal appears without primary antibody. | Possible endogenous detection activity (general IHC practice). | Review the detection chemistry and its blocking controls (general IHC practice). |
Experiment evidence
IHC & IF images showing ID1 staining
M00945 · IHCImmunohistochemical analysis of paraffin-embedded human kidney, using Id1 Antibody.
M00945 · IFImmunofluorescent analysis of Hela cells, using Id1 Antibody .
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000125968-ID1/tissue/cerebral+cortex#imid_7768640
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000125968-ID1/tissue/colon#imid_7768566
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000125968-ID1/tissue/liver#imid_7768559
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000125968-ID1/tissue/kidney#imid_7768568
HPA · ICC-IF PC-3Human Protein Atlas immunofluorescence, PC-3, antibody HPA027874; green = ID1, blue = nucleus, red = microtubules.
HPA · ICC-IF PC-3Human Protein Atlas immunofluorescence, PC-3, antibody HPA027874; green = ID1, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for ID1 IHC & IF
🧪Run bronchus first: respiratory epithelial cells should stain (HPA: High in respiratory epithelial cells). Use adipose tissue as the negative tissue, focusing on adipocytes (HPA: Not detected in adipocytes); on the bronchus slide, neighboring cells without specific chromogen should retain their counterstain, but the supplied HPA rows do not establish which neighboring cell types lack ID1.
Positive control tissue: Bronchus (Respiratory epithelial cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show ID1 in PC-3, U-251MG, U2OS, A-549, HeLa, with annotated localisation: Nucleoplasm (enhanced) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control; an isotype control matched to the primary antibody’s host species and immunoglobulin class (or nonimmune IgG for a polyclonal primary); and ID1-knockout material or a validated peptide-block control (standard IHC practice). On bronchus sections, quench endogenous peroxidase and inspect mucus and inflammatory cells for background chromogen (standard IHC practice).
⚠️Feasibility: No target-specific fixation window or fixation effect is reported, and the selected M00945 paraffin-section caption does not state a fixative (selected-SKU caption: fixative not stated). Antigen-retrieval dependency is unreported, so optimize retrieval empirically for paraffin IHC; assess nuclear staining with a counterstain while allowing for cytoplasmic localization (UniProt P41134: nucleus and cytoplasm; HPA subcellular: enhanced nucleoplasmic localization). The supplied evidence does not establish whether frozen sections or IF are easier; bronchial mucus and endogenous peroxidase in inflammatory cells warrant background checks (standard IHC practice).
HPA tissue IHC evidence for ID1
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Pending RNA-based expert annotation.). 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 | Respiratory epithelial cells | High | Protein (IHC) | HPA → |
| Pancreas | Exocrine glandular cells | High | Protein (IHC) | HPA → |
| Placenta | Trophoblastic cells | High | Protein (IHC) | HPA → |
| Soft tissue | Chondrocytes | High | Protein (IHC) | HPA → |
| Tonsil | Non-germinal center cells | High | Protein (IHC) | HPA → |
Section 3
Advanced ID1 IHC Tips
Troubleshoot ID1 staining in paraffin sections by checking retrieval, compartment pattern, cell identity, and controls before assigning biological meaning.
How should I retrieve ID1 in paraffin sections when nuclear staining is weak?
Use Tris-EDTA at pH 9.0 for heat-induced retrieval at 95–98 °C for 20 min (page retrieval rule: nuclear antigen). Let sections cool consistently, then compare staining with a matched section processed in the same run (standard IHC practice). ID1 is found in both nucleus and cytoplasm, so assess both compartments while checking whether the expected nuclear signal becomes clearer (UniProt P41134: subcellular location; HPA tissue IHC: cytoplasmic and nuclear expression). If staining remains weak, test a citrate buffer at pH 6.0 as a fallback on adjacent sections (standard IHC practice). Judge any change against background and tissue morphology before adopting it (standard IHC practice).
Could fixation explain inconsistent ID1 staining across paraffin specimens?
Target-specific ID1 sensitivity to fixation is unknown from the supplied evidence (supplied fixation evidence: none). The selected M00945 image shows staining in paraffin-embedded human kidney, but its caption does not state the fixative (M00945 tissue-IHC caption). Record the fixative, fixation duration, processing history, and section age for each specimen before comparing staining intensity (standard IHC practice). Run specimens with similar processing together and include a control section in each run so changes in retrieval or detection can be distinguished from specimen variation (standard IHC practice). Do not assign a particular fixation effect to ID1 without a controlled comparison using the same antibody and detection workflow (supplied fixation evidence: none).
Should a valid ID1 stain appear in nuclei, cytoplasm, or both?
Assess nuclear and cytoplasmic signal separately because ID1 is annotated in both compartments (UniProt P41134: subcellular location). Tissue IHC also reports cytoplasmic and nuclear expression across tissues, whereas the subcellular image summary emphasizes nucleoplasm (HPA tissue IHC: profile; HPA subcellular: nucleoplasm). Check that nuclear staining follows intact nuclei under the counterstain and that cytoplasmic signal stays within recognizable cell boundaries (standard IHC practice). If only diffuse extracellular color appears, inspect section folds, tissue edges, and background before calling it ID1 (standard IHC practice). Record the compartment alongside intensity because a single combined score could hide a change in the staining pattern (standard IHC practice).
How can antibody epitope coverage affect interpretation of ID1 staining?
ID1 has 2 annotated isoforms, ID-A and ID-B, and its bHLH domain spans residues 53–105 (UniProt P41134: isoforms and domains). The supplied evidence does not specify the catalog antibody’s epitope or establish whether it recognizes both isoforms (supplied antibody evidence: M00945 tissue-IHC caption). Check the antibody’s documented immunogen or epitope before treating a negative section as absence of every ID1 isoform (standard IHC practice; UniProt P41134: isoforms). The record lists no signal peptide, transmembrane segment, glycosylation site, or modified residue, so those features provide no basis here for assigning an unusual staining pattern to processing (UniProt P41134: topology, processing, and modifications). Compare staining with an independent antibody if isoform coverage matters to the conclusion (standard IHC practice).
How should I investigate ID1 localisation by multiplex IF?
Treat IF as a separate application and confirm that the antibody performs under the chosen IF conditions before interpreting a merged image (standard IF practice; M00945 tissue-IHC caption: paraffin tissue IHC). Multiplex ID1 with a marker for the expected cell population, selecting that marker from the specimen’s cell identities rather than assuming every positive nucleus belongs to one lineage (standard IF practice; HPA tissue IHC: cell-specific observations). Choose a fluorophore channel that separates ID1 signal from the specimen’s autofluorescence, and inspect single-channel controls (standard IF practice). ID1 has no transmembrane segment and is reported in nucleus and cytoplasm, so use permeabilisation that allows antibody access to intracellular epitopes (UniProt P41134: topology and subcellular location; standard IF practice). Compare nuclear signal with a nuclear counterstain and keep exposure settings consistent (standard IF practice).
What should I check when ID1 chromogenic staining has high background?
First inspect a no-primary control and the counterstained section for diffuse color, edge staining, or deposits that do not follow cell boundaries (standard IHC practice). Apply the peroxidase block required by the detection workflow and check for endogenous enzyme signal before interpreting DAB color (standard chromogenic IHC practice). Review blocking, wash steps, antibody concentration, and detection time one variable at a time while keeping the retrieval condition fixed (standard IHC practice; page retrieval rule: nuclear antigen). ID1 can be nuclear and cytoplasmic, so cytoplasmic staining alone is not automatically background (UniProt P41134: subcellular location; HPA tissue IHC: profile). Require a reproducible cellular pattern and a quiet no-primary control before scoring weak signal (standard IHC practice).
How should I score ID1 across sections with different cell composition? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, then record the percentage of positive cells and staining intensity within that population (standard IHC practice; UniProt P41134: subcellular location). An H-score can combine intensity categories from 0–3 with their percentages to give a value from 0–300 (standard IHC practice). If spatial distribution matters, count positive cells per mm² of viable tissue and report the area sampled (standard IHC practice). Normalize comparisons to the number of eligible cells or the viable tissue area, using the same thresholds and imaging conditions across sections (standard IHC practice). Report nuclear and cytoplasmic results separately so differences in localisation remain visible (standard IHC practice; HPA tissue IHC: profile).
How can I distinguish true ID1 positivity from staining artefact?
Look for signal within intact nuclei or cytoplasm and assess it in an identified cell population, since ID1 is reported in both compartments (UniProt P41134: subcellular location; HPA tissue IHC: profile). HPA reports high staining in bronchial respiratory epithelial cells and pancreatic exocrine glandular cells, providing documented patterns for comparison when those tissues are examined (HPA tissue IHC: positive cell observations). Treat color concentrated at section edges, folds, or necrotic areas as suspect until a well-preserved region shows the same cellular pattern (standard IHC practice). Check a no-primary control and peroxidase block for endogenous enzyme signal before accepting weak DAB staining (standard chromogenic IHC practice). Interpret unexpected positive cells cautiously because the HPA tissue IHC reliability is Approved with RNA-based expert annotation pending (HPA tissue IHC: reliability).
Boster reagents
Best ID1 / DNA-binding protein inhibitor ID-1 IHC Antibodies
M00945 has IHC data from paraffin-embedded human kidney and IF data from Hela cells (catalog image captions). Its listed reactivity covers human, mouse, and rat (catalog reactivity).
Real IHC data
Anti-Id1 Rabbit Monoclonal Antibody
Cat # M00945
M00945 is shown in IHC on paraffin-embedded human kidney (IHC image caption). M00945 is also shown in IF on Hela cells and lists IHC, ICC, and IF applications with human, mouse, and rat reactivity (IF image caption; catalog applications and reactivity).
Which to pick: Choose M00945 for paraffin-section tissue IHC based on its human kidney image; the fixative is unreported (IHC image caption). Choose M00945 for IF/ICC because it is a rabbit monoclonal with an IF image from Hela cells and lists both ICC and IF (catalog host and clonality; IF image caption; catalog applications). For mouse or rat samples, M00945 lists reactivity with those species, while its shown IHC tissue data are human (catalog reactivity; IHC image caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P41134 (ID1_HUMAN, DNA-binding protein inhibitor ID-1).
- Human Protein Atlas. ID1 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. ID1 subcellular location (ICC-IF): Localized to the nucleoplasm..
- Human Protein Atlas. ID1 antibody validation summary (4 antibodies).
- Bone marrow and peripheral blood expression of ID1 in human gastric carcinoma patients is a bona fide indicator of lymph node and peritoneal metastasis. British journal of cancer 2009 — PMC2714249.
- Id Proteins Contribute to Tumor Development and Metastatic Colonization in a Model of Bladder Carcinogenesis. Bladder cancer (Amsterdam, Netherlands) 2015 — PMC4927902.
- Id1 and Id3 co-expression correlates with clinical outcome in stage III-N2 non-small cell lung cancer patients treated with definitive chemoradiotherapy. Journal of translational medicine 2013 — PMC3567999.
- PubMed PMID:8086456 — UniProt-cited evidence.
- PubMed PMID:8294468 — UniProt-cited evidence.
- PubMed PMID:7637581 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for ID1 (DNA-binding protein inhibitor ID-1). Expected localisation Cytoplasmic and nuclear tissue staining (HPA tissue IHC). Antigen retrieval: Tris-EDTA pH 9.0 HIER, 95–98 °C, 20 min (rule: nuclear antigen). Validated IHC antibody: M00945 at 1:50 (the product's own IHC validation image is shown). Controls (HPA): positive Bronchus (Respiratory epithelial cells); negative Adipose tissue. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Paraffin-embedded human kidney tissue; fixative not specified (datasheet M00945); 2. Fixation: Image fixative and duration unreported (datasheet M00945); 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 Tris-EDTA buffer, pH 9.0 , 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 monoclonal (clone ACC-9) anti-ID1, 1:50 (datasheet M00945); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation conditions consistent across sections. (standard IHC practice; not target-specific); Caveat: Adipocytes may be unstained despite broad tissue expression (HPA tissue IHC); Isoform / epitope: 2 isoforms (ID-A/ID-B); epitope coverage unknown (UniProt); Regulation: No expression stimulus specified (UniProt). Evidence: UniProt P41134, HPA tissue IHC · Approved, HPA ICC-IF · nucleoplasm., Datasheet M00945, PMC2714249, PMC4927902. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/ID1/immunohistochemistry-ihc-guide.