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ID1 / DNA-binding protein inhibitor ID-1 · IHC design guide
Design Immunohistochemistry for ID1
Plan chromogenic ID1 IHC in paraffin sections using respiratory epithelial cells and trophoblasts as high-staining references (HPA tissue IHC). Assess both nuclear and cytoplasmic signal (HPA tissue IHC) with matched controls.
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 | Cytoplasm and nucleus (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 paraffin-section fixation consistent across controls. (selected-SKU IHC image M00945) | |
| Caveat | Adipocytes and ovarian stroma may show no signal (HPA tissue IHC) | |
| Regulation | Intensity regulation unreported (UniProt) | |
| Isoform / epitope | 2 isoforms, ID-A and ID-B; epitope effect unresolved (UniProt) |
Section 1
Recommended ID1 IHC & IF Protocols
The catalog antibody’s IHC-P protocol is accompanied by published ID1 staining protocols from three articles (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 Tris-EDTA pH 9.0 retrieval at 95–98 °C for 20 min (page antigen retrieval); try citrate retrieval when following PMC2714249.
Section 2
What Is the Expected ID1 Staining Pattern?
ID1 is reported in the nucleus and cytoplasm, with no transmembrane segment (UniProt P41134). In tissue sections, HPA reports cytoplasmic and nuclear staining across tissues, including high staining in bronchial respiratory epithelium, pancreatic exocrine cells and placental trophoblasts (HPA tissue IHC). HPA rates its tissue IHC profile Approved, with RNA-based expert annotation pending (HPA tissue IHC).
| Nuclear and cytoplasmic staining in respiratory epithelial cells, with clear cell outlines and little background. | This matches the reported compartments and a high-staining cell population (UniProt P41134; HPA tissue IHC). Judge intensity against nearby tissue and controls: HPA's High category describes its observed pattern, not a required intensity for every section (HPA tissue IHC; general IHC practice). |
| Staining appears mainly as a sharp cell-surface rim or extracellular deposit. | That distribution conflicts with ID1's nuclear and cytoplasmic localization and lack of a transmembrane segment (UniProt P41134). Consider nonspecific deposition or detection artefact; a membrane-like pattern alone is weak evidence for ID1 (UniProt P41134; general IHC practice). |
| Strong signal appears in adipocytes or ovarian stromal cells. | HPA reports ID1 as not detected in those specific cell populations (HPA tissue IHC). Review cell identification, controls and possible cross-reactivity or endogenous detection activity before interpreting the signal; this comparison does not make the entire tissue a negative control (HPA tissue IHC; general IHC practice). |
| Color spreads across stroma, lumina or tissue edges without distinct stained cells. | Diffuse deposit obscures the nuclear and cytoplasmic pattern needed for interpretation (UniProt P41134; general IHC practice). Compare a no-primary control and inspect washing, blocking and detection background before assigning cell-level ID1 staining (general IHC practice). |
| A bronchus, pancreas or placenta section shows no convincing signal in its listed high-staining cells. | That differs from HPA's observations for respiratory epithelial, exocrine glandular or trophoblastic cells, respectively (HPA tissue IHC). First confirm the expected cells are present, then assess controls and staining conditions; one negative section does not overturn the reported tissue pattern (HPA tissue IHC; general IHC practice). |
💡Expected ID1 appearanceA convincing positive shows nuclear and cytoplasmic signal in identifiable cells such as HPA-listed high-staining respiratory epithelium, while isolated membrane rims or cell-free deposits warrant scrutiny (UniProt P41134; HPA tissue IHC; general IHC practice).
| Cell population selected for assessment | HPA lists high staining in bronchial respiratory epithelium, pancreatic exocrine cells, trophoblasts, chondrocytes and tonsillar non-germinal-center cells; it lists adipocytes and ovarian stromal cells as not detected (HPA tissue IHC). Assess named cells rather than treating an entire organ as uniformly positive or negative. |
| Compartment evidence by application | UniProt lists nucleus and cytoplasm, and HPA tissue IHC reports both (UniProt P41134; HPA tissue IHC). HPA ICC-IF instead reports enhanced nucleoplasmic localization (HPA subcellular). The IF result supports nuclear localization but does not set an IHC staining requirement. |
| Antibody validation scope | The supplied antibody list marks CAB025915 Approved for IHC and three separate antibodies Enhanced for ICC, with no IHC status listed for those three (HPA antibodies). Keep application-specific validation distinct; the supplied record does not establish Enhanced IHC validation. |
| Isoforms and epitope coverage | UniProt lists ID-A and ID-B and a bHLH domain at residues 53–105 (UniProt P41134). The payload gives no antibody epitope or isoform coverage, so neither the observed pattern nor a weak signal can be assigned to one isoform from these records. |
| Antigen retrieval choice | Retrieval conditions can be compared with suitable controls during paraffin-section assay setup (general IHC practice). No supplied source reports ID1-specific retrieval or fixation sensitivity; HPA staining levels and UniProt topology cannot establish such an effect. |
| Endogenous chromogenic activity | Endogenous activity can add color independently of primary-antibody binding in a chromogenic assay (general IHC practice). Check detection and no-primary controls when unexpected cell types or diffuse deposits stain; this is a workflow consideration, not an ID1-specific finding. |
| Situation | Likely cause | Next action |
|---|---|---|
| Expected high-staining cells are present, but the section is blank. | The assay may have failed, or that specimen may differ from the HPA observation; the supplied sources do not identify an ID1-specific fixation cause (HPA tissue IHC; general IHC practice). | Confirm morphology and cell identity, then check a concurrently stained positive section and detection controls. Review the IHC-validated antibody's actual instructions before changing retrieval or dilution (general IHC practice). |
| Signal is restricted to a membrane-like rim. | A rim-only pattern conflicts with the reported nucleus and cytoplasm and the absence of a transmembrane segment (UniProt P41134). | Inspect cell boundaries and a no-primary control, and compare the compartment pattern in a listed high-staining cell population before scoring the rim as ID1 (HPA tissue IHC; general IHC practice). |
| Adipocytes or ovarian stromal cells stain strongly. | HPA lists those cells as not detected; cross-reactivity, endogenous detection activity or misidentified cells are possibilities, not proven causes (HPA tissue IHC; general IHC practice). | Verify the cell type on the counterstained section and compare no-primary and detection controls. Report the discrepancy if it persists (general IHC practice). |
| Color covers tissue edges or extracellular spaces. | A deposit without defined stained cells does not reproduce the reported intracellular pattern (UniProt P41134; HPA tissue IHC). Background from the staining workflow is possible (general IHC practice). | Compare a no-primary section, inspect washing and blocking, and score only interpretable cells once background is controlled (general IHC practice). |
| Nuclear signal is clear, but cytoplasmic signal is weak. | HPA ICC-IF reports enhanced nucleoplasmic localization, while tissue IHC reports both compartments; the records give no fixed nuclear-to-cytoplasmic intensity ratio (HPA subcellular; HPA tissue IHC). | Record the compartments actually visible and compare appropriate controls and listed positive cells. Do not require equal compartment intensity without assay-specific evidence (HPA tissue IHC; general IHC practice). |
| IF/ICC shows nucleoplasmic staining: is that expected? | Yes. HPA reports enhanced nucleoplasmic localization in ICC-IF, consistent with UniProt's nuclear localization (HPA subcellular; UniProt P41134). | Interpret that result using the separate IF/ICC guide and its controls; the ICC-IF antibody ratings do not establish an IHC protocol or Enhanced IHC validation (HPA antibodies; 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 comparator, focusing on adipocytes (HPA: Not detected in adipocytes). On the bronchus slide, cells without specific signal should show only counterstain; the supplied HPA row does not identify a validated negative cell type within bronchus (HPA: High in respiratory epithelial cells).
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 clonality; and ID1 knockout material as a biological negative. Block endogenous peroxidase for chromogenic IHC and check for background associated with bronchial mucus (standard IHC practice).
⚠️Feasibility: A target-specific fixation window and retrieval dependency are unreported in the supplied evidence; optimize retrieval empirically (selected-SKU caption: paraffin-embedded human kidney; fixative not stated). The selected-SKU caption does not report a fixative (selected-SKU caption: fixative not stated). The evidence does not establish whether frozen sections or IF are easier; ICC-IF images show nucleoplasmic localisation, which can guide localisation assessment (HPA subcellular: nucleoplasm, enhanced).
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 section IHC by checking retrieval, compartment, controls and cell level scoring before interpreting signal (UniProt P41134; HPA tissue IHC).
How should I retrieve ID1 in paraffin sections when nuclear staining is weak?
Start with Tris-EDTA at pH 9.0, heated to 95–98 °C for 20 min (page retrieval setting). Let sections cool in retrieval buffer, then compare staining with a matched section processed identically except for retrieval (standard IHC practice). Assess both nuclear and cytoplasmic signal, because ID1 has been reported in both compartments (UniProt P41134; HPA tissue IHC). If signal remains weak, compare a shorter heating time or an alternative retrieval buffer on serial sections, changing one condition at a time (standard IHC optimization). Judge improvement against tissue morphology and background as well as signal intensity (standard IHC practice).
Could fixation explain variable ID1 staining between paraffin blocks?
ID1 specific sensitivity to fixation is unknown from the supplied evidence; the catalog antibody caption identifies paraffin embedded human kidney but does not state the fixative (M00945 tissue-IHC caption). Record each block’s fixative, fixation duration and processing history before attributing a staining difference to biology (standard IHC practice). If those histories differ, stain serial sections in the same run with the same retrieval setting and detection conditions (standard IHC practice; page retrieval setting). Check whether weak signal tracks a particular processing history while morphology and control staining remain interpretable (standard IHC practice). Do not infer fixation tolerance from tissue expression patterns or ID1’s lack of a transmembrane segment and annotated glycosylation (HPA tissue IHC; UniProt P41134).
Should convincing ID1 staining be nuclear, cytoplasmic or both?
Evaluate nuclear and cytoplasmic staining separately: both are reported for ID1 in tissue IHC (HPA tissue IHC), while enhanced nucleoplasmic localisation is reported in ICC/IF (HPA subcellular). Its role in regulating transcription supports examining the nuclear compartment closely, but does not make every cytoplasmic signal erroneous (UniProt P41134). Use a nuclear counterstain to confirm that apparent nuclear chromogen lies within intact nuclei (standard IHC practice). Compare compartment patterns in morphologically intact cells and across serial sections processed together (standard IHC practice). If staining is confined to tissue edges or damaged regions, investigate processing and detection before assigning an ID1 localisation (standard IHC practice).
Can this stain distinguish ID1 isoforms or reveal an epitope problem?
The record lists 2 ID1 isoforms, ID-A and ID-B, and places the bHLH domain at residues 53–105 (UniProt P41134). No epitope location or isoform recognition profile is supplied for the catalog antibody, so a positive stain cannot be assigned to one isoform (supplied antibody evidence). Check the antibody’s documented immunogen and epitope before using staining differences to make an isoform claim (standard antibody validation practice). If staining changes after retrieval, compare serial sections under matched detection conditions; altered epitope accessibility is a possible technical explanation (standard IHC practice). Confirm any proposed isoform difference with an independently validated isoform specific method (standard antibody validation practice).
How can IF help verify which cells and compartments carry ID1 signal?
For a separate IF experiment, multiplex ID1 with a marker validated for the expected cell type, then inspect each channel and the merged image (standard IF practice). Choose fluorophores after checking the specimen’s autofluorescence, favoring a spectrally separated channel with low background for ID1 (standard IF practice). Because ID1 is reported in the nucleoplasm and cytoplasm and has no transmembrane segment, use a mild permeabilisation condition that permits access to intracellular epitopes (HPA subcellular; UniProt P41134; standard IF practice). Include single stain and secondary only controls to assess bleed through and nonspecific fluorescence (standard IF practice). An IF result does not establish identical staining behavior in paraffin section chromogenic IHC (standard assay validation practice).
What should I check when ID1 chromogen appears diffuse or widespread?
First compare the stained section with a no primary control and inspect where chromogen accumulates (standard IHC practice). Block endogenous peroxidase before peroxidase based detection, and assess whether DAB signal follows tissue edges, folds or damaged areas (standard chromogenic IHC practice). Optimize antibody concentration and incubation only with an appropriate positive control in the same run; no dilution is supplied for this antibody (standard IHC practice; supplied antibody evidence). Widespread cellular staining alone is inconclusive because tissue IHC reports cytoplasmic and nuclear ID1 expression across tissues (HPA tissue IHC). Require coherent cell boundaries, interpretable nuclei and low control background before calling diffuse staining positive (standard IHC practice).
How should I score ID1 when nuclear and cytoplasmic staining differ? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and scoring compartment before counting, then report nuclear and cytoplasmic results separately (standard IHC quantification practice; HPA tissue IHC). For each compartment, record the percentage of positive cells and an intensity category; an H-score can combine those measurements consistently across specimens (standard IHC quantification practice). If spatial abundance is the question, count positive cells per mm² of viable tissue instead (standard IHC quantification practice). Normalize cell counts to the measured viable area, or positive fractions to the number of eligible cells in the same compartment and region (standard IHC quantification practice). Apply one threshold and identical exclusion rules to all specimens (standard IHC quantification practice).
How do I distinguish credible ID1 positivity from staining artefact?
A credible result has staining within intact cells, with nuclear or cytoplasmic localisation compatible with reported ID1 patterns (UniProt P41134; HPA tissue IHC). Compare the implicated cell population with reference patterns: respiratory epithelial cells, exocrine glandular cells and trophoblastic cells have reported high staining (HPA tissue IHC). Treat positivity in an unexpected cell population as a finding to verify, since those reference patterns do not exclude other ID1 expressing cells (HPA tissue IHC). Discount signal restricted to section edges, folds or necrotic tissue, and check a no primary control for endogenous enzyme or detection background (standard IHC practice). Require reproducibility across sections before interpreting a compartment change biologically (standard IHC practice).
Boster reagents
Best ID1 / DNA-binding protein inhibitor ID-1 IHC Antibodies
M00945 has IHC data from paraffin-embedded human kidney (M00945 IHC image caption) and IF data from Hela cells (M00945 IF image caption); listed reactivity covers human, mouse, and rat (catalog reactivity).
Real IHC data
Anti-Id1 Rabbit Monoclonal Antibody
Cat # M00945
M00945 is listed for IHC, ICC, and IF in human, mouse, and rat (catalog applications and reactivity). Its images show IHC in paraffin-embedded human kidney and IF in Hela cells (M00945 image captions).
Which to pick: Choose M00945 for paraffin-section IHC: its own image shows staining in human kidney (M00945 IHC image caption). The fixative is unreported (M00945 IHC image caption). For IF/ICC and human, mouse, or rat samples, M00945 is the listed rabbit monoclonal option, with an IF image from Hela cells (catalog host, clone, applications, and reactivity; M00945 IF 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 Cytoplasm and nucleus (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 paraffin-section fixation consistent across controls. (selected-SKU IHC image M00945); Caveat: Adipocytes and ovarian stroma may show no signal (HPA tissue IHC); Isoform / epitope: 2 isoforms, ID-A and ID-B; epitope effect unresolved (UniProt); Regulation: Intensity regulation unreported (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.