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DIDO1 / Death-inducer obliterator 1 · IHC design guide
Design Immunohistochemistry for DIDO1
Plan chromogenic DIDO1 IHC in paraffin sections using the catalog antibody’s 1:100–1:300 dilution range (datasheet A06019). Assess the mainly nuclear tissue pattern (HPA tissue IHC) with consistent fixation and cell-type scoring (standard IHC practice).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
DIDO1 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 | Mainly nuclear in tissue (HPA tissue IHC); cytoplasmic location annotated (UniProt) | |
| Staining pattern | Mainly nuclear staining in most tissues (HPA tissue IHC) | |
| Antigen retrieval | Tris-EDTA pH 9.0 HIER, heat-mediated (datasheet A06019) | |
| Positive control | Adrenal gland+4 more · see all | |
| Negative control | None in HPA (detected in all 45 tissues); use no-primary + isotype controls |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent across sections (standard IHC practice; not target-specific) | |
| Caveat | Antibody staining and RNA show medium consistency (HPA tissue IHC) | |
| Regulation | No specific induction reported (UniProt) | |
| Isoform / epitope | Four isoforms; verify antibody epitope coverage (UniProt) |
Section 1
Recommended DIDO1 IHC & IF Protocols
The catalog antibody protocol and two published DIDO1 IHC methods cover paraffin sections (datasheet A06019; PMC12994943; PMC8188684).
| Sample | Paraffin-embedded human tonsil tissue; fixative not specified (datasheet A06019) |
| Fixation | Image fixative and duration unreported (datasheet A06019); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat retrieval: Tris-EDTA pH 9.0 (datasheet A06019); 20 min, 95–100 °C (standard) |
| 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-DIDO1, 1:100-1:300 (datasheet A06019) |
| 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 | DIDO1-positive staining in glandular cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: Mainly nuclear expression in most tissues. No signal in the no-primary control. |
💡Decision noteStart with Tris-EDTA pH 9.0 heat retrieval for the catalog antibody (datasheet A06019); use citrate pH 6.0 when reproducing either published method (PMC12994943; PMC8188684).
Section 2
What Is the Expected DIDO1 Staining Pattern?
DIDO1 should stain mainly nuclei across many tissues (HPA tissue IHC: mainly nuclear expression; UniProt Q9BTC0: ubiquitous). Examples of cells with high staining include bone marrow hematopoietic cells, adrenal glandular cells and bronchial respiratory epithelial cells (HPA tissue IHC). HPA rates the tissue pattern Supported, with medium antibody–RNA consistency and external verification pending (HPA tissue IHC). DIDO1 has no transmembrane segment (UniProt Q9BTC0 topology).
| Predominant nuclear chromogenic signal in expected cells, including adrenal glandular or bone marrow hematopoietic cells. | This matches the reported pattern and high staining in those cell populations (HPA tissue IHC). Assess compartment and cell identity together; a dark deposit alone does not establish specific DIDO1 staining (general IHC practice). |
| Signal is exclusively membranous, or strongly cytoplasmic while nuclei remain unstained. | A purely membranous pattern conflicts with DIDO1 topology (UniProt Q9BTC0 topology). Cytoplasmic DIDO1 is possible (UniProt Q9BTC0 subcellular location), but a cytoplasm-only tissue pattern conflicts with HPA’s mainly nuclear IHC profile; investigate artefact or nonspecific binding (HPA tissue IHC; general IHC practice). |
| Prominent staining appears in cells outside the reported positive populations within a reference tissue. | HPA identifies specific strongly stained cell populations, but supplies no negative-cell list (HPA tissue IHC). Treat the finding as unresolved: cross-reactivity or endogenous chromogenic activity is possible, and an unlisted cell type cannot automatically be called DIDO1-negative (general IHC practice). |
| Chromogen forms a diffuse haze over cells or extracellular areas, obscuring nuclei. | The distribution cannot be scored confidently against the mainly nuclear reference pattern (HPA tissue IHC). Review background controls, blocking, antibody concentration and detection steps to locate nonspecific signal (general IHC practice). |
| No convincing signal appears in an expected positive population, such as bone marrow hematopoietic cells. | This disagrees with the reported high level for that population (HPA tissue IHC). First check section quality, the staining run and positive control; a failed stain alone does not establish biological absence (general IHC practice). |
💡Expected DIDO1 appearanceCall a section positive when identifiable cells show predominantly nuclear chromogenic staining, with high signal expected in HPA-listed populations; diffuse haze or isolated membrane staining warrants review (HPA tissue IHC; UniProt Q9BTC0 topology; general IHC practice).
| Compartment and cell identity | HPA reports mainly nuclear tissue staining and high levels in selected cell populations (HPA tissue IHC). Score nuclei in identified cells rather than treating all tissue staining as equivalent (general IHC practice). |
| Context-dependent localization | UniProt places DIDO1 in nucleus and cytoplasm and describes spindle translocation in early mitosis after loss of H3K4me3 interaction (UniProt Q9BTC0). This allows a context-dependent exception; it does not redefine the predominant tissue IHC pattern (HPA tissue IHC). |
| Tissue evidence limits | HPA lists no negative or low-staining tissues in the supplied record and rates the pattern Supported, with external verification pending (HPA tissue IHC). Use listed positives as references without declaring unlisted tissues negative. |
| Antibody evidence | HPA049904 and CAB004374 each have IHC status Supported; each has ICC status Uncertain (HPA antibodies). Their IHC status supports comparison with the tissue pattern, but does not establish identical staining for every antibody or assay. |
| Isoforms and processing | UniProt lists 4 isoforms and a single 1–2240 chain, with no signal peptide or propeptide (UniProt Q9BTC0). The supplied record does not map the IHC antibody epitope, so isoform-specific staining cannot be predicted. |
| IF/ICC Q: what localization is reported? | A: HPA approves nucleoplasm as the main ICC-IF location, with vesicles and cytosol also reported (HPA subcellular). This is a localization cross-check; the listed antibodies’ ICC validation is Uncertain (HPA antibodies). |
| Situation | Likely cause | Next action |
|---|---|---|
| Known-positive tissue is blank. | The result conflicts with HPA’s high staining in the specified positive cells (HPA tissue IHC); an unsuccessful staining run is possible (general IHC practice). | Verify the positive-control section, reagent sequence, retrieval and detection controls before interpreting absence of DIDO1 (general IHC practice). |
| Nuclei are obscured by widespread background. | Diffuse deposit prevents comparison with the mainly nuclear pattern (HPA tissue IHC); excess antibody or incomplete blocking may contribute (general IHC practice). | Inspect a negative reagent control, then review blocking, washes, dilution and chromogen development (general IHC practice). |
| Only cell borders stain. | An exclusively membranous pattern is inconsistent with the lack of a transmembrane segment (UniProt Q9BTC0 topology). | Compare with a known-positive nuclear pattern and check negative reagent controls for nonspecific detection (HPA tissue IHC; general IHC practice). |
| Unexpected cells stain strongly. | HPA supplies selected positive populations but no negative-cell list (HPA tissue IHC); cross-reactivity or endogenous detection activity may mimic signal (general IHC practice). | Identify the stained cells, check reagent controls and compare the compartment with a listed positive population before assigning specificity (general IHC practice; HPA tissue IHC). |
| A few dividing cells show signal near spindle structures. | DIDO1 can move to the mitotic spindle during early mitosis under the stated chromatin-binding condition (UniProt Q9BTC0). | Assess whether staining is confined to plausible mitotic cells; keep predominant nuclear staining as the tissue-level reference (UniProt Q9BTC0; HPA tissue IHC; general IHC practice). |
| IF/ICC appears more cytosolic than the tissue IHC section. | HPA reports additional cytosol and vesicle localization in ICC-IF, while tissue IHC is mainly nuclear (HPA subcellular; HPA tissue IHC). | Compare compartment patterns within each assay and note that the listed antibodies have ICC status Uncertain (HPA antibodies). |
Experiment evidence
IHC & IF images showing DIDO1 staining
A06019 · IHCImmunohistochemical analysis of paraffin-embedded human tonsil. 1, Antibody was diluted at 1:200 (4° overnight). 2, Tris-EDTA, pH9.0 was use
A06019S186 · IHCImmunohistochemistry (IHC) analyzes of Dio-1 (S186) pAb in paraffin-embedded human breast carcinoma tissue at 1:50.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000101191-DIDO1/tissue/cerebral+cortex#imid_1613288
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000101191-DIDO1/tissue/colon#imid_1613222
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000101191-DIDO1/tissue/liver#imid_15508210
HPA · PancreasHuman Protein Atlas IHC, Pancreas. Source: https://www.proteinatlas.org/ENSG00000101191-DIDO1/tissue/pancreas#imid_1613174
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody CAB004374; green = DIDO1, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody CAB004374; green = DIDO1, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for DIDO1 IHC & IF
🧪Run bone marrow first: hematopoietic cells should stain (HPA: High in hematopoietic cells). HPA detects DIDO1 in all 45 scored tissues, so there is no supported negative tissue; no-primary and isotype controls carry the negative, while enucleated erythrocytes on the positive slide should lack nuclear staining but are not validated DIDO1-negative cells (HPA: no negative tissue rows; standard IHC practice).
Positive control tissue: Adrenal gland (Glandular cells, HPA High)
Negative control tissue: None in HPA: DIDO1 is detected in all 45 scored tissues. Use a no-primary (secondary-only) and an isotype control instead.
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show DIDO1 in A-431, U-251MG, U2OS, HeLa, with annotated localisation: Nucleoplasm (approved) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only slide and a control matched to the primary antibody’s host and immunoglobulin class if monoclonal, or host-matched nonimmune IgG if polyclonal; use DIDO1 knockout material or a validated peptide block as a biological negative (standard IHC practice). Quench endogenous peroxidase in bone marrow before chromogenic detection (standard IHC practice).
⚠️Feasibility: The selected paraffin-section caption reports Tris-EDTA retrieval at pH 9.0, but its fixative is unreported, and no target-specific fixation window or fixation effect is supplied (catalog antibody A06019 caption). IF/ICC localization has supporting images in A-431, U-251MG, U2OS and HeLa cells (HPA: subcellular images); the supplied evidence does not establish that frozen sections are easier. Bone marrow’s endogenous peroxidase can create chromogenic background (standard IHC practice).
HPA tissue IHC evidence for DIDO1
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Supported — Medium 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 |
|---|---|---|---|---|
| None in HPA: DIDO1 is detected in all 45 scored tissues. Use a no-primary (secondary-only) and an isotype control instead. | ||||
Section 3
Advanced DIDO1 IHC Tips
Troubleshoot DIDO1 staining in paraffin-section chromogenic IHC using the catalog antibody’s tonsil example and compartment-specific controls.
What retrieval should I try first for weak DIDO1 staining?
Start with heat-mediated Tris-EDTA retrieval at pH 9.0 for paraffin-section DIDO1 IHC (datasheet A06019). The catalog antibody’s human tonsil image used that retrieval, a 1:200 primary dilution at 4°C overnight, and a 1:200 secondary for 45 minutes at room temperature (datasheet A06019). Compare weak slides with a tonsil control processed in the same run, and score nuclear staining separately from diffuse background (datasheet A06019; HPA tissue IHC: mainly nuclear). If signal remains weak, test an alternative retrieval buffer as a controlled fallback, changing only that step and checking tissue integrity (general IHC practice).
Could fixation be causing inconsistent nuclear staining?
DIDO1-specific sensitivity to fixation is unknown: the catalog tonsil caption describes paraffin embedding but does not state the fixative (datasheet A06019). Record the fixative, fixation duration, section age, and retrieval conditions for each specimen before attributing differences to DIDO1 biology (general IHC practice). Process a reference section alongside each batch using Tris-EDTA at pH 9.0 and the catalog’s 1:200 primary condition, then compare nuclear signal and tissue preservation (datasheet A06019; general IHC practice). If fixation histories differ, interpret staining comparisons cautiously; HPA nuclear patterns do not establish how this antibody responds to fixation (HPA tissue IHC: mainly nuclear).
How should I evaluate nuclear, cytoplasmic, or spindle-like DIDO1 staining?
Expect predominantly nuclear staining across many tissues, while recognizing reported nucleoplasmic, cytosolic, and vesicular locations (HPA tissue IHC: mainly nuclear; HPA subcellular: nucleoplasm, cytosol, vesicles). DIDO1 can move toward the nucleus after pro-apoptotic stimuli and toward the mitotic spindle during early mitosis, but these annotations do not establish what a given paraffin section must show (UniProt Q9BTC0). Score nuclear and cytoplasmic compartments separately in at least 2 matched sections when investigating a shift (general IHC practice). Examine mitotic morphology and neighboring cells before calling isolated spindle-like chromogen a biological redistribution (UniProt Q9BTC0; general IHC practice).
Can this stain distinguish DIDO1 isoforms or modified epitopes?
DIDO1 has 4 annotated isoforms and multiple modified residues, including phosphoserines at positions 60 and 114 (UniProt Q9BTC0). The supplied catalog caption gives staining conditions but no epitope map or isoform-reactivity data, so a positive section cannot identify a particular isoform or phosphorylation state (datasheet A06019; UniProt Q9BTC0). Compare the antibody’s documented immunogen or epitope with isoform sequences before assigning molecular specificity, and retain a common reference section for staining comparisons (general IHC practice). If an experimental perturbation changes staining, confirm the proposed isoform or modification with an independently validated, modification-aware method before interpreting the IHC result (general IHC practice).
How can IF help check an ambiguous IHC compartment pattern?
Use IF as a separate validation experiment for an ambiguous chromogenic pattern; the supplied catalog evidence documents paraffin-section IHC conditions, not IF performance (datasheet A06019). Multiplex DIDO1 with a validated marker for the expected cell population, such as glandular cells in appendix, and include single-stain controls to assess channel bleed-through (HPA tissue IHC: high in appendix glandular cells; general IF practice). Choose a red or far-red fluorophore after checking tissue autofluorescence in each channel (general IF practice). Because DIDO1 lacks a transmembrane segment and is reported in intracellular compartments, select permeabilisation for the intracellular epitope and verify that it preserves nuclear detail (UniProt Q9BTC0; general IF practice).
What should I check when DIDO1 chromogen covers whole sections?
First compare a no-primary control with the DIDO1 slide to identify secondary-reagent, endogenous-enzyme, or chromogen background (general chromogenic IHC practice). Apply a peroxidase block before DAB detection and optimize protein blocking and washing for the tissue; these are general workflow steps, not DIDO1-specific findings (general chromogenic IHC practice). The catalog tonsil example used 1:200 primary overnight at 4°C and 1:200 secondary for 45 minutes; use those conditions as a documented comparison point (datasheet A06019). If diffuse staining persists, titrate antibody concentration while checking that mainly nuclear signal remains distinguishable from cytoplasmic haze (HPA tissue IHC: mainly nuclear; general IHC practice).
How should I quantify DIDO1 across heterogeneous tissue sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, because DIDO1 tissue IHC is mainly nuclear and reported expression spans many tissues (HPA tissue IHC: mainly nuclear; UniProt Q9BTC0: ubiquitous). For nuclear staining, record percent positive cells and intensity on a 0–3 scale, then calculate an H-score from 0–300 using a fixed threshold (general IHC practice). Normalize counts to the number of evaluable cells in the same annotated region; use positive-cell density per mm² when region area is the comparison unit (general IHC practice). Exclude folds, necrosis, and edge artefacts consistently, and report nuclear and cytoplasmic results separately rather than combining their signals (general IHC practice).
When is an apparent DIDO1-positive cell likely an artefact?
Prioritize convincing nuclear staining in intact cells, since tissue IHC is mainly nuclear, while allowing for documented cytosolic and vesicular localization (HPA tissue IHC: mainly nuclear; HPA subcellular: nucleoplasm, cytosol, vesicles). Check cell identity against the annotated region: HPA reports high staining in appendix glandular cells and bone-marrow hematopoietic cells, but these examples do not validate every stained cell (HPA tissue IHC). Staining restricted to section edges, folds, or necrotic areas warrants repeat assessment with a matched reference section (general IHC practice). Compare a no-primary control and peroxidase-blocked slide before interpreting broad DAB deposition as DIDO1, and review at least 2 intact fields (general chromogenic IHC practice).
Boster reagents
Best DIDO1 / Death-inducer obliterator 1 IHC Antibodies
IHC images document paraffin-embedded human tonsil and breast carcinoma (image captions: A06019, A06019S186); catalog reactivity also lists mouse and rat (catalog: reactivity). No IF image is supplied (catalog: IF image captions).
Real IHC data
Anti-Dio-1 DIDO1 Antibody
Cat # A06019
Real IHC data
Anti-Dio-1 (S186) DIDO1 Antibody
Cat # A06019S186
A06019 will render with human tonsil paraffin-section IHC at 1:200; its listed applications include IF, and its reactivity is human and mouse (A06019 image caption; catalog: applications and reactivity). A06019S186 will render with human breast carcinoma paraffin-section IHC at 1:50; its listed reactivity is human, mouse and rat (A06019S186 image caption; catalog: reactivity).
Which to pick: For tissue IHC, choose A06019 for the documented human tonsil example or A06019S186 for the documented human breast carcinoma example; both captions specify paraffin-embedded tissue, and neither reports the fixative (respective IHC image captions). For IF/ICC, A06019-1 lists both applications and is described as polyclonal, but has no supplied IF image; A06019 lists IF and 1:50 without an IF image (catalog: applications, dilution, clonality and IF image captions). For rat tissue IHC, A06019S186 lists rat reactivity and IHC, while its own IHC image shows human tissue only (catalog: reactivity and applications; A06019S186 image caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q9BTC0 (DIDO1_HUMAN, Death-inducer obliterator 1).
- Human Protein Atlas. DIDO1 tissue IHC expression (reliability: Supported).
- Human Protein Atlas. DIDO1 subcellular location (ICC-IF): Mainly localized to the nucleoplasm. In addition localized to vesicles and cytosol..
- Human Protein Atlas. DIDO1 antibody validation summary (2 antibodies).
- CSE1L, DIDO1 and RBM39 in colorectal adenoma to carcinoma progression. Cellular oncology (Dordrecht, Netherlands) 2012 — PMC12994943.
- Serum anti-DIDO1, anti-CPSF2, and anti-FOXJ2 antibodies as predictive risk markers for acute ischemic stroke. BMC medicine 2021 — PMC8188684.
- PubMed PMID:16127461 — UniProt-cited evidence.
- PubMed PMID:9205841 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for DIDO1 (Death-inducer obliterator 1). Expected localisation Mainly nuclear in tissue (HPA tissue IHC); cytoplasmic location annotated (UniProt). Antigen retrieval: Tris-EDTA pH 9.0 HIER, heat-mediated (datasheet A06019). Validated IHC antibody: A06019 at 1:100-1:300 (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Glandular cells); negative None in HPA (detected in all 45 tissues); use no-primary + isotype controls. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Paraffin-embedded human tonsil tissue; fixative not specified (datasheet A06019); 2. Fixation: Image fixative and duration unreported (datasheet A06019); 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 retrieval: Tris-EDTA pH 9.0 (datasheet A06019); 20 min, 95–100 °C (standard); 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-DIDO1, 1:100-1:300 (datasheet A06019); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across sections (standard IHC practice; not target-specific); Caveat: Antibody staining and RNA show medium consistency (HPA tissue IHC); Isoform / epitope: Four isoforms; verify antibody epitope coverage (UniProt); Regulation: No specific induction reported (UniProt). Evidence: UniProt Q9BTC0, HPA tissue IHC · Supported, HPA ICC-IF · Mainly localized to the nucleoplasm. In addition localized to…, Datasheet A06019, PMC12994943, PMC8188684. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/DIDO1/immunohistochemistry-ihc-guide.