This website uses cookies to ensure you get the best experience on our website.
- Table of Contents
DHODH / Dihydroorotate dehydrogenase (quinone), mitochondrial · IHC design guide
Design Immunohistochemistry for DHODH
Plan paraffin-section DHODH IHC with the IHC-validated antibody M04035-1 (datasheet M04035-1). Compare cytoplasmic staining in high-staining glandular cells with caudate glial cells reported as undetected (HPA tissue IHC), and score the stained cell populations.
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
DHODH 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 stain (HPA tissue IHC); inner mitochondrial membrane (UniProt) | |
| Staining pattern | Cytoplasmic; high in glandular, endocrine and lymphoid tissues (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet M04035-1) | |
| Positive control | Appendix+4 more · see all | |
| Negative control | Caudate |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent across sections (standard IHC practice; not target-specific); Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image M04035-1) | |
| Caveat | Staining has low agreement with RNA expression (HPA tissue IHC) | |
| Regulation | No expression regulation annotated (UniProt) | |
| Isoform / epitope | No annotated isoforms or cleavage; epitope map unknown (UniProt) |
Section 1
Recommended DHODH IHC & IF Protocols
The catalog antibody protocol is accompanied by 2 published DHODH IHC protocols: adult human brain (PMC6814620) and hepatocellular carcinoma tissue (PMC11910144).
| Sample | Paraffin-embedded human breast cancer tissue; fixative not specified (datasheet M04035-1) |
| Fixation | Image fixative and duration unreported (datasheet M04035-1); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat retrieval: EDTA pH 8.0 (datasheet M04035-1); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet M04035-1) |
| Primary antibody | Mouse monoclonal (clone 2G7) anti-DHODH, 2 μg/ml (datasheet M04035-1) |
| Primary incubation | Overnight at 4 °C (datasheet M04035-1) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet M04035-1) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | DHODH-positive staining in glandular cells of appendix (HPA tissue IHC: High). HPA tissue profile: General cytoplasmic expression. High levels in glandular epithelia, endocrine tissues and in lymphoid tissues. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA pH 8.0 retrieval (datasheet M04035-1); adjust retrieval for the sample using the published conditions (PMC6814620; PMC11910144).
Section 2
What Is the Expected DHODH Staining Pattern?
DHODH is anchored in the mitochondrial inner membrane by residues 11–30, so paraffin-section IHC should chiefly show cytoplasmic staining in expressing cells (UniProt Q02127 topology; HPA tissue IHC). HPA reports high staining in glandular epithelia, kidney tubules and Purkinje cells (HPA tissue IHC). Its tissue IHC reliability is Approved, with low agreement between staining and RNA data and external verification pending (HPA tissue IHC).
| Cytoplasmic stain is strong in appendix or colon glandular cells, or kidney tubule cells (HPA tissue IHC). | This matches reported high-staining cells (HPA tissue IHC). A mitochondrial distribution is consistent with inner-membrane anchoring, although chromogenic IHC alone cannot resolve that membrane (UniProt Q02127 topology; standard IHC practice). |
| Staining is predominantly nuclear, with little cytoplasmic signal. | Recheck localisation and controls: a nuclear-dominant IHC result diverges from HPA's general cytoplasmic tissue pattern (HPA tissue IHC). Some nucleoplasmic signal is reported in ICC-IF, so nuclear signal alone does not prove artefact (HPA subcellular ICC-IF). |
| Unexpected cells stain strongly while the expected positive cells do not. | Consider nonspecific antibody binding or endogenous chromogen-generating activity (standard IHC practice). Compare cells within the same section against HPA's cell-level pattern before calling the unexpected signal DHODH (HPA tissue IHC). |
| Brown colour spreads evenly across cells or tissue, obscuring cellular boundaries. | A uniform haze is hard to assign to DHODH's reported cytoplasmic pattern (HPA tissue IHC). Check reagent-only and no-primary controls for background from the detection workflow (standard IHC practice). |
| Known high-staining glandular cells or kidney tubules show no signal (HPA tissue IHC). | First suspect an assay or sample problem; absence in one section cannot establish that DHODH is absent (standard IHC practice). HPA's low staining–RNA agreement also limits tissue-level inference from a single IHC result (HPA tissue IHC). |
💡Expected DHODH appearanceA convincing positive is predominantly cytoplasmic staining in HPA high-staining cells, such as colon glandular cells or kidney tubules; isolated nuclear-dominant or uniform background colour needs control-based review (HPA tissue IHC; standard IHC practice).
| Topology and compartment | Residues 11–30 anchor DHODH in the mitochondrial inner membrane; residues 31–395 face the intermembrane space (UniProt Q02127 topology). Interpret routine chromogenic IHC at the cellular-compartment level (standard IHC practice). |
| Tissue and cell selection | HPA reports high staining in appendix, breast, colon, duodenum, endometrium and gallbladder glandular cells, kidney tubules, and cerebellar Purkinje cells (HPA tissue IHC). Caudate glial cells were not detected (HPA tissue IHC). |
| Strength of IHC evidence | HPA010123 has Approved IHC status; HPA011942 has no IHC status in the supplied antibody record (HPA antibodies). HPA also flags low staining–RNA agreement and pending external verification (HPA tissue IHC). |
| RNA versus protein readout | HPA calls liver RNA tissue-enriched, but that designation does not supply a liver IHC staining level (HPA tissue IHC). Choose protein-positive control cells from the reported IHC observations (HPA tissue IHC). |
| Q: What should IF/ICC show? | A: Mainly mitochondrial signal; HPA additionally reports nucleoplasmic and cytosolic localisation (HPA subcellular ICC-IF). Its ICC-IF images include MCF-7, U2OS and A-549; these observations do not specify a paraffin-IHC protocol (HPA subcellular ICC-IF). |
| Situation | Likely cause | Next action |
|---|---|---|
| No stain in an expected high-staining cell population (HPA tissue IHC). | The staining run, detection reagents or tissue preservation may have failed (standard IHC practice); this record gives no DHODH-specific fixation sensitivity. | Check the run with a reported high-staining tissue and review antigen retrieval, antibody dilution and detection steps against the chosen validated IHC procedure (HPA tissue IHC; standard IHC practice). |
| Widespread brown precipitate or diffuse tissue colour. | Endogenous detection activity, excess chromogen development or nonspecific background may contribute (standard IHC practice). | Inspect no-primary and reagent-only controls; then review endogenous-activity blocking, washes and development time (standard IHC practice). Judge any remaining cellular stain against HPA's cytoplasmic pattern (HPA tissue IHC). |
| A strong nuclear-dominant IHC pattern replaces expected cytoplasmic stain (HPA tissue IHC). | Nonspecific staining is possible (standard IHC practice), although HPA reports additional nucleoplasmic localisation by ICC-IF (HPA subcellular ICC-IF). | Check controls and whether cytoplasmic staining remains in HPA high-staining cells; avoid scoring nuclear-only IHC as confirmed DHODH from this evidence (HPA tissue IHC; standard IHC practice). |
| Strong signal appears in cells outside the expected high-staining population. | Cross-reactivity or endogenous detection activity is possible (standard IHC practice); HPA's cell-level observations are a comparison, not an exhaustive list of expressing cells (HPA tissue IHC). | Compare adjacent cell types and controls, then review the antibody's IHC validation before assigning the unexpected signal to DHODH (HPA antibodies; standard IHC practice). |
| Signal is faint in a low-staining comparison population. | HPA reports low staining in oral mucosa squamous epithelium and several other specified cell populations (HPA tissue IHC). | Confirm that a reported high-staining control works before increasing signal development; compare like cell types rather than whole-tissue averages (HPA tissue IHC; standard IHC practice). |
| IF/ICC and chromogenic IHC appear to disagree on subcellular detail. | HPA describes mitochondrial plus additional nucleoplasmic and cytosolic ICC-IF signal, while tissue IHC is described broadly as cytoplasmic (HPA subcellular ICC-IF; HPA tissue IHC). | Score each assay against its own reported pattern and controls; do not require paraffin-section chromogen to resolve the inner mitochondrial membrane (UniProt Q02127 topology; standard IHC practice). |
Experiment evidence
IHC & IF images showing DHODH staining
M04035-1 · IHCIHC analysis of DHODH using anti-DHODH antibody (M04035-1). DHODH was detected in a paraffin-embedded section of human breast cancer tissue.
M04035-1 · IHCIHC analysis of DHODH using anti-DHODH antibody (M04035-1). DHODH was detected in a paraffin-embedded section of human gastric cancer tissue
PB10056 · IHCIHC analysis of DHODH using anti-DHODH antibody (PB10056). DHODH was detected in a paraffin-embedded section of mouse intestine tissue. Heat
PB10056 · IHCIHC analysis of DHODH using anti-DHODH antibody (PB10056). DHODH was detected in a paraffin-embedded section of human lung cancer tissue. He
M04035 · IFIF analysis of DHODH using anti-DHODH antibody (M04035). DHODH was detected in an immunocytochemical section of MCF-7 cells. Enzyme antigen
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000102967-DHODH/tissue/cerebral+cortex#imid_4198436
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000102967-DHODH/tissue/colon#imid_4198370
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000102967-DHODH/tissue/liver#imid_4198371
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000102967-DHODH/tissue/kidney#imid_4198364
HPA · ICC-IF MCF-7Human Protein Atlas immunofluorescence, MCF-7, antibody HPA010123; green = DHODH, blue = nucleus, red = microtubules.
HPA · ICC-IF MCF-7Human Protein Atlas immunofluorescence, MCF-7, antibody HPA010123; green = DHODH, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for DHODH IHC & IF
🧪Run breast first and require staining in glandular cells (HPA: High in breast glandular cells). Use caudate glial cells as the negative tissue comparison (HPA: Not detected in caudate glial cells); on the breast slide, assess adjacent non-glandular cells for background, without assuming they are DHODH-negative.
Positive control tissue: Appendix (Glandular cells, HPA High)
Negative control tissue: Caudate (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show DHODH in MCF-7, U2OS, A-549, with annotated localisation: Mitochondria (enhanced) (HPA subcellular).
Technical controls: Include a no-primary (secondary-only) section, a concentration-matched nonimmune mouse IgG isotype control matched to the primary antibody’s subclass, and DHODH knockout material as a biological negative (selected-SKU caption: mouse anti-DHODH antibody). For breast sections, check endogenous peroxidase and biotin background because the reported detection uses a streptavidin–biotin complex with DAB (selected-SKU tissue-IHC caption).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the exact selected-SKU tissue-IHC caption does not state a fixative. Paraffin-section IHC is documented with heat-mediated EDTA retrieval at pH 8.0; the caption does not establish how staining depends on retrieval (selected-SKU tissue-IHC caption). Frozen-section IHC or tissue IF cannot be judged easier from the supplied evidence; breast staining with the reported biotin-based detection should be checked for endogenous biotin background (selected-SKU tissue-IHC caption).
HPA tissue IHC evidence for DHODH
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 |
|---|---|---|---|---|
| Appendix | Glandular cells | High | Protein (IHC) | HPA → |
| Breast | Glandular cells | High | Protein (IHC) | HPA → |
| Cerebellum | Purkinje cells | High | Protein (IHC) | HPA → |
| Colon | Glandular cells | High | Protein (IHC) | HPA → |
| Duodenum | Glandular cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Caudate | Glial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced DHODH IHC Tips
Troubleshoot DHODH staining in paraffin sections by checking retrieval, compartment, cellular context and controls before interpreting chromogenic signal.
What retrieval should I start with if DHODH staining is weak?
Start with heat-mediated retrieval in EDTA at pH 8.0 (datasheet M04035-1). The catalog antibody detected DHODH in a paraffin-embedded human breast cancer section after this retrieval, using 2 μg/mL primary antibody overnight at 4°C (datasheet M04035-1). If staining is weak, compare a longer heating or cooling interval on adjacent sections while keeping antibody concentration and detection conditions fixed (standard IHC practice). Include a known positive tissue and a no-primary control in each comparison so a stronger DAB signal can be judged against background (HPA: high in breast glandular cells; standard IHC practice).
Could fixation explain variable DHODH staining between paraffin blocks?
DHODH-specific fixation sensitivity is unknown: the selected tissue-IHC caption identifies a paraffin-embedded section but does not state its fixative (datasheet M04035-1). Compare blocks with documented fixation histories using the same pH 8.0 EDTA retrieval, primary concentration and DAB development conditions (datasheet M04035-1; standard IHC practice). Look for differences in staining alongside differences in tissue preservation, and avoid treating a faint block as biologically negative without an internal or matched positive control (standard IHC practice). Record fixative, fixation duration and processing history for each block; the supplied evidence does not establish a DHODH-specific optimum for any of them (datasheet M04035-1).
Where should convincing DHODH chromogenic staining appear?
Expect a predominantly cytoplasmic pattern compatible with mitochondria, since DHODH is assigned to the mitochondrial inner membrane and tissue IHC shows general cytoplasmic expression (UniProt Q02127 topology; HPA tissue IHC). Its 11–30 membrane segment anchors the protein, while residues 31–395 face the intermembrane space; routine DAB microscopy will not resolve those membrane surfaces (UniProt Q02127 topology; standard IHC practice). Compare cellular staining with tissue architecture rather than calling every diffuse cytoplasmic deposit mitochondrial (standard IHC practice). Nuclear or broadly diffuse signal deserves control checks, although independent subcellular imaging reports additional nucleoplasmic and cytosolic localisation (HPA subcellular: enhanced mitochondrial, approved additional locations).
How do topology and epitope uncertainty affect DHODH staining?
The supplied record lists one 395-residue DHODH chain, no annotated isoforms, and no annotated glycosylation or modified-residue sites (UniProt Q02127). Its membrane-spanning region is residues 11–30, with residues 31–395 on the intermembrane side of the inner mitochondrial membrane (UniProt Q02127 topology). The supplied caption does not identify the catalog antibody's epitope, so a retrieval failure cannot be assigned to a particular residue or modification (datasheet M04035-1). If staining varies, compare retrieval and primary-antibody conditions on adjacent sections, then check whether the resulting cellular pattern remains compatible with mitochondrial DHODH (standard IHC practice; UniProt Q02127 localisation).
How should I investigate DHODH localisation by multiplex IF?
Use a separate IF validation series before translating this paraffin-section DAB result to fluorescence, because the selected caption documents chromogenic tissue IHC (datasheet M04035-1). Pair DHODH with a glandular epithelial marker such as cytokeratin when examining glandular regions, and include a mitochondrial marker to assess its expected organelle pattern (HPA tissue IHC: high in glandular cells; HPA subcellular: mainly mitochondrial; standard IF practice). Choose a far-red fluorophore after measuring tissue autofluorescence in unstained sections, and run single-stain controls for channel separation (standard IF practice). Titrate permeabilisation to give antibody access beyond the plasma and outer mitochondrial membranes to the intermembrane-facing DHODH region, while checking that mitochondrial morphology remains intact (UniProt Q02127 topology; standard IF practice).
How can I distinguish DHODH signal from DAB background?
The selected assay used 10% goat serum blocking, a biotinylated goat anti-mouse secondary, streptavidin–biotin detection and DAB (datasheet M04035-1). For chromogenic IHC, include a peroxidase block and no-primary control, then inspect any DAB signal produced without the primary antibody (standard IHC practice). If widespread background remains, titrate the 2 μg/mL primary concentration and secondary detection separately while keeping retrieval at pH 8.0 initially (datasheet M04035-1; standard IHC practice). Check section edges, damaged tissue and biotin-related background before assigning granular staining to DHODH (standard IHC practice).
How should DHODH staining be scored across tissue regions? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and region of interest before scoring, because DHODH staining varies across tissue cell types (HPA tissue IHC). For glandular cells, record the percentage of positive cells and an H-score using intensity categories 0–3; use positive-cell density per mm² when cell counts differ substantially between regions (standard IHC practice). Normalise each score to the relevant viable-cell count or analysed viable-tissue area, and exclude necrosis, folds and section edges using a prespecified rule (standard IHC practice). Keep retrieval, illumination, DAB development and scoring thresholds consistent across slides, with a shared positive control to monitor run variation (standard IHC practice).
What findings would make a DHODH-positive result credible?
A credible result shows cellular staining compatible with DHODH's mainly mitochondrial location and fits the sampled cell population, with clean no-primary and detection controls (UniProt Q02127; HPA subcellular; standard IHC practice). High staining in glandular cells is plausible, whereas an isolated strong signal in a population reported as undetected, such as caudate glial cells, needs closer validation (HPA tissue IHC). Examine nuclear-only staining, section edges, necrotic zones and deposits in the no-primary control before interpreting DAB as DHODH (HPA subcellular: additional nucleoplasmic location; standard IHC practice). Treat concordance with the HPA tissue pattern as supportive rather than definitive, because its tissue-IHC entry reports low agreement with RNA expression and pending external verification (HPA tissue IHC: Approved reliability description).
Boster reagents
Best DHODH / Dihydroorotate dehydrogenase (quinone), mitochondrial IHC Antibodies
Anti-DHODH antibodies have IHC images from human breast cancer and mouse intestine paraffin sections, plus an IF/ICC image from MCF-7 cells (catalog image captions).
Real IHC data
Anti-DHODH Antibody ® (monoclonal, 2G7)
Cat # M04035-1
Real IHC data
Anti-DHODH Antibody ®
Cat # PB10056
Real IF data
Anti-DHODH Antibody ® (monoclonal, 4E3)
Cat # M04035
M04035-1 is IHC-listed and shown on human breast cancer paraffin sections; PB10056 is IHC-listed and shown on mouse intestine paraffin sections (catalog applications; respective image captions). M04035 is IF/ICC-listed and shown in MCF-7 cells (catalog applications; M04035 image caption).
Which to pick: For human tissue IHC, choose mouse monoclonal M04035-1: its own paraffin-section caption documents EDTA pH 8 retrieval, while the fixative is unreported (catalog antibody details; M04035-1 image caption). For IF/ICC, choose mouse monoclonal M04035, which has an MCF-7 cell image (catalog applications; M04035 image caption). For IHC across human, mouse and rat, consider rabbit PB10056: its IHC reactivity lists all three species, and its own mouse intestine paraffin-section caption documents EDTA pH 8 retrieval; the fixative is unreported (PB10056 catalog reactivity; PB10056 image caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q02127 (PYRD_HUMAN, Dihydroorotate dehydrogenase (quinone), mitochondrial).
- Human Protein Atlas. DHODH tissue IHC expression (reliability: Approved).
- Human Protein Atlas. DHODH subcellular location (ICC-IF): Mainly localized to the mitochondria. In addition localized to the nucleoplasm and cytosol..
- Human Protein Atlas. DHODH antibody validation summary (2 antibodies).
- Metabolomics profiles delineate uridine deficiency contributes to mitochondria-mediated apoptosis induced by celastrol in human acute promyelocytic leukemia cells. Oncotarget 2016 — PMC5216817.
- Brain pyrimidine nucleotide synthesis and Alzheimer disease. Aging 2019 — PMC6814620.
- Designer Exosomes for Targeted Delivery of a Novel Therapeutic Cargo to Enhance Sorafenib-Mediated Ferroptosis in Hepatocellular Carcinoma. Frontiers in oncology 2022 — PMC9263838.
- Targeting YBX1-m5C mediates RNF115 mRNA circularisation and translation to enhance vulnerability of ferroptosis in hepatocellular carcinoma. Clinical and translational medicine 2025 — PMC11910144.
- PubMed PMID:1446837 — UniProt-cited evidence.
- PubMed PMID:7487077 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for DHODH (Dihydroorotate dehydrogenase (quinone), mitochondrial). Expected localisation Cytoplasmic stain (HPA tissue IHC); inner mitochondrial membrane (UniProt). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet M04035-1). Validated IHC antibody: M04035-1 at 2 μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Appendix (Glandular cells); negative Caudate. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human breast cancer tissue; fixative not specified (datasheet M04035-1); 2. Fixation: Image fixative and duration unreported (datasheet M04035-1); 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: EDTA pH 8.0 (datasheet M04035-1); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet M04035-1); 8. Primary antibody: Mouse monoclonal (clone 2G7) anti-DHODH, 2 μg/ml (datasheet M04035-1); 9. Primary incubation: Overnight at 4 °C (datasheet M04035-1). Watch for: Fixation: Keep fixation consistent across sections (standard IHC practice; not target-specific); Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image M04035-1); Caveat: Staining has low agreement with RNA expression (HPA tissue IHC); Isoform / epitope: No annotated isoforms or cleavage; epitope map unknown (UniProt); Regulation: No expression regulation annotated (UniProt). Evidence: UniProt Q02127, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the mitochondria. In addition localized to the…, Datasheet M04035-1, PMC6814620, PMC11910144. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/DHODH/immunohistochemistry-ihc-guide.