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CYGB / Cytoglobin · IHC design guide
Design Immunohistochemistry for CYGB
Plan paraffin-section CYGB IHC around the predominantly cytoplasmic tissue pattern (HPA tissue IHC). Use skeletal-muscle myocytes as a high-staining reference and interpret intensity with the reported medium consistency between staining and RNA (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
CYGB 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 | Predominantly cytoplasmic in tissue IHC (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic staining, prominent in some immune cells (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6 HIER, heat-mediated (datasheet A01197) | |
| Positive control | Skeletal muscle+4 more · see all | |
| Negative control | Bone marrow+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent across sections (standard IHC practice; not target-specific) | |
| Caveat | Staining and RNA show only medium consistency (HPA tissue IHC) | |
| Regulation | Expression regulation is unreported (UniProt) | |
| Isoform / epitope | No listed isoforms; one 1–190 chain (UniProt) |
Section 1
Recommended CYGB IHC & IF Protocols
The catalog antibody protocol is paired with published CYGB IHC methods for human liver and glioma sections (PMC5849339; PMC3663650).
| Sample | Paraffin-embedded mouse intestine tissues; fixative not specified (datasheet A01197) |
| Fixation | Image fixative and duration unreported (datasheet A01197); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat retrieval: Citrate pH 6, 20 min (datasheet A01197) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A01197) |
| Primary antibody | Rabbit anti-CYGB, 0.5-1μg/ml (datasheet A01197) |
| Primary incubation | Overnight at 4 °C (datasheet A01197) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet A01197) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | CYGB-positive staining in myocytes of skeletal muscle (HPA tissue IHC: High). HPA tissue profile: General cytoplasmic expression, most abundant in a subset of immune cells. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated citrate retrieval at pH 6 (datasheet A01197); the glioma protocol also reports citrate retrieval (PMC3663650).
Section 2
What Is the Expected CYGB Staining Pattern?
CYGB can localize to the cytoplasm and nucleus and has no transmembrane segment (UniProt Q8WWM9). In tissue IHC, expect predominantly cytoplasmic staining, especially in a subset of immune cells (HPA tissue IHC). Skeletal muscle myocytes are reported as high, while several glandular and neuronal cell populations are medium (HPA tissue IHC). The tissue profile is Approved, with medium consistency between antibody staining and RNA expression (HPA tissue IHC).
| Strong cytoplasmic staining in skeletal muscle myocytes, with weaker or variable staining in other cells. | This fits the reported high myocyte signal and general cytoplasmic tissue pattern (HPA tissue IHC). Compare intensity within identified cell types; a uniformly dark section does not reproduce this cell-specific profile (HPA tissue IHC). |
| Signal lies mainly on cell borders, in extracellular material, or as a surface rim. | Treat this as a localization mismatch requiring investigation: CYGB has no transmembrane segment, and the reported tissue pattern is cytoplasmic (UniProt Q8WWM9; HPA tissue IHC). Check morphology and detection controls before assigning the signal to CYGB (general IHC practice). |
| Strong staining appears in a cell population reported as undetected, such as skin fibroblasts. | This conflicts with the reference cell-level observation (HPA tissue IHC: skin fibroblasts not detected). Cross-reactivity or endogenous detection activity are possibilities, not diagnoses; assess matched controls and whether the stained cells were identified correctly (general IHC practice). |
| Color spreads across cells, stroma, and blank areas without clear cellular boundaries. | A diffuse deposit cannot establish CYGB-positive cells; the reference describes cellular, predominantly cytoplasmic staining (HPA tissue IHC). Review reagent background, blocking, washing, and the no-primary control (general IHC practice). |
| No staining is visible in skeletal muscle myocytes on an otherwise interpretable section. | The expected high signal is absent (HPA tissue IHC: skeletal muscle myocytes High). First check tissue identity and assay controls, then review retrieval, primary incubation, and detection as general IHC troubleshooting steps; the sources do not establish a CYGB-specific retrieval requirement. |
💡Expected CYGB appearanceCall a result positive when identifiable cells show predominantly cytoplasmic signal, with strong skeletal muscle myocyte staining (HPA tissue IHC: High); widespread cell-free deposit or a membrane-only rim is suspect (HPA tissue IHC; UniProt Q8WWM9 topology).
| Cell-level reference | HPA reports high skeletal muscle myocytes and medium bronchial respiratory epithelial, colon glandular, and cerebral cortical neuronal cells (HPA tissue IHC). Select a comparator by its named cell population, since a tissue contains multiple cell types (general IHC practice). |
| Negative comparator | HPA reports no detection in bone marrow hematopoietic cells, skin fibroblasts, and prostate glandular cells (HPA tissue IHC). These are cell-specific observations, not a promise that every cell or structure in those sections will be unstained. |
| Evidence strength | The tissue profile is Approved with medium staining–RNA consistency (HPA tissue IHC). HPA017757 has IHC Approved status; HPA061642 has no listed IHC status (HPA antibodies). Treat an unexpected pattern as needing controls rather than as definitive biology. |
| Protein features | CYGB is a single 1–190 chain with no signal peptide, propeptide, or transmembrane segment (UniProt Q8WWM9). Those features support an intracellular expectation; they do not identify an antibody epitope or establish antigen retrieval conditions. |
| IF/ICC Q: Is a nuclear-speckle signal compatible with CYGB? | A: Yes in the reported ICC-IF images: nuclear speckles are the supported main location, with additional cytosol staining (HPA subcellular). UniProt also lists nucleus and cytoplasm (UniProt Q8WWM9). Interpret that observation in its IF/ICC context; it does not replace the predominantly cytoplasmic tissue-IHC reference (HPA tissue IHC). |
| Detection chemistry | Endogenous activity can contribute chromogenic staining, depending on the detection system (general IHC practice). Use controls appropriate to the chosen detection chemistry; neither HPA nor UniProt supplies a CYGB-specific interference claim. |
| Situation | Likely cause | Next action |
|---|---|---|
| The expected skeletal muscle myocyte signal is absent. | The section may lack evaluable myocytes, or an assay step may have failed; HPA reports high myocyte staining (HPA tissue IHC). | Confirm cell identity and a working positive control, then review retrieval, primary incubation, and detection records (general IHC practice). Do not infer a CYGB-specific fixation effect from this result. |
| Most of the section has diffuse chromogen. | Background from the detection workflow may obscure the cellular pattern (general IHC practice). HPA describes general cytoplasmic expression with enrichment in a subset of immune cells (HPA tissue IHC). | Inspect a no-primary control and review blocking, washes, and chromogen development (general IHC practice). Score CYGB only where cells and their cytoplasm can be resolved. |
| Skin fibroblasts or bone marrow hematopoietic cells stain strongly. | The result differs from HPA's not-detected observations for those cells (HPA tissue IHC). Cell misidentification, cross-reactivity, or endogenous detection activity may explain it (general IHC practice). | Recheck morphology and matched controls; if signal persists, seek independent antibody or orthogonal evidence before calling those cells CYGB-positive (general IHC practice). |
| Signal forms a membrane rim or coats extracellular structures. | That pattern is difficult to reconcile with CYGB's lack of a transmembrane segment and the cytoplasmic tissue profile (UniProt Q8WWM9; HPA tissue IHC). | Check the no-primary control and inspect the deposit against cell boundaries (general IHC practice). Do not score a rim alone as the expected CYGB pattern. |
| A nuclear signal accompanies cytoplasmic staining. | Nuclear localization is listed by UniProt, and nuclear speckles are supported in HPA ICC-IF (UniProt Q8WWM9; HPA subcellular). The tissue-IHC profile emphasizes cytoplasm (HPA tissue IHC). | Record nuclear and cytoplasmic compartments separately. Compare the tissue signal with cell identity and controls before deciding whether the nuclear component is specific (general IHC practice). |
| Heart muscle RNA enrichment seems inconsistent with a slide's staining intensity. | HPA lists tissue-enhanced RNA in heart muscle but only medium consistency between antibody staining and RNA expression (HPA tissue IHC). RNA abundance does not specify the staining intensity of each cell (general IHC interpretation). | Score the observed cell-level IHC pattern and document the discrepancy. Avoid converting RNA enrichment into an expected chromogenic intensity for an individual cell (HPA tissue IHC). |
Experiment evidence
IHC & IF images showing CYGB staining
A01197 · IHCIHC analysis of Cytoglobin using anti-Cytoglobin antibody (A01197). Cytoglobin was detected in paraffin-embedded section of mouse intestine
A01197 · IHCIHC analysis of Cytoglobin using anti-Cytoglobin antibody (A01197). Cytoglobin was detected in paraffin-embedded section of mouse cardiac mu
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000161544-CYGB/tissue/cerebral+cortex#imid_4658304
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000161544-CYGB/tissue/colon#imid_4658246
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000161544-CYGB/tissue/liver#imid_4658247
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000161544-CYGB/tissue/kidney#imid_4658240
HPA · ICC-IF Hep-G2Human Protein Atlas immunofluorescence, Hep-G2, antibody HPA017757; green = CYGB, blue = nucleus, red = microtubules.
HPA · ICC-IF Hep-G2Human Protein Atlas immunofluorescence, Hep-G2, antibody HPA017757; green = CYGB, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for CYGB IHC & IF
🧪Run skeletal muscle first and score myocytes for CYGB staining (HPA: High in skeletal-muscle myocytes). Use bone marrow hematopoietic cells as the negative tissue (HPA: Not detected); on the positive slide, assess morphologically distinct nonmyocytes as internal negative candidates and confirm that they show only background-level chromogen before treating them as controls.
Positive control tissue: Skeletal muscle (Myocytes, HPA High)
Negative control tissue: Bone marrow (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show CYGB in Hep-G2, SH-SY5Y, U-251MG, with annotated localisation: Nuclear speckles (supported) (HPA subcellular).
Technical controls: Include no-primary (secondary-only) and host-matched rabbit IgG isotype controls (caption: rabbit primary antibody), plus CYGB-knockout material or a validated peptide-block control. In skeletal muscle, check for endogenous peroxidase signal and tissue pigment with the no-primary control before interpreting DAB staining (caption: DAB detection).
⚠️Feasibility: No target-specific fixation window or fixation effect is reported, and the selected A01197 paraffin-section caption does not state a fixative (caption: fixative unreported). The caption uses heat retrieval in citrate buffer at pH 6 for 20 minutes, but does not establish that retrieval is required under other conditions (caption: retrieval conditions). Frozen sections have no demonstrated ease advantage in the supplied evidence; IF/ICC has supported nuclear-speckle and cytosolic localization, while skeletal-muscle pigment or endogenous peroxidase can complicate chromogenic scoring (HPA: subcellular localization; caption: DAB detection).
HPA tissue IHC evidence for CYGB
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Medium consistency between antibody staining and RNA expression data.). 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 |
|---|---|---|---|---|
| Skeletal muscle | Myocytes | High | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | Medium | Protein (IHC) | HPA → |
| Appendix | Glandular cells | Medium | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | Medium | Protein (IHC) | HPA → |
| Cerebellum | Cells in molecular layer | Medium | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Bone marrow | Hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Caudate | Glial cells | Not detected | Protein (IHC) | HPA → |
| Cervix | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Liver | Cholangiocytes | Not detected | Protein (IHC) | HPA → |
| Lymph node | Germinal center cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced CYGB IHC Tips
Troubleshoot CYGB staining in paraffin sections by checking retrieval, cell identity, subcellular pattern and matched controls before scoring.
What retrieval should I start with if CYGB staining is weak?
Start with heat-mediated retrieval in citrate buffer at pH 6 for 20 minutes (datasheet A01197). The selected paraffin-section image used this condition before incubation with 1 μg/mL antibody overnight at 4°C (datasheet A01197). If staining remains weak, compare a modestly longer heating period on adjacent sections while keeping the antibody concentration and detection steps constant (standard IHC practice). Include a section processed under the documented condition in each comparison so a gain in signal can be judged against any increase in background (standard IHC practice). Do not interpret stronger staining alone as better retrieval without checking the expected cytoplasmic pattern (HPA tissue IHC).
Could fixation explain faint or uneven CYGB staining?
Target-specific fixation sensitivity is unknown, and the selected paraffin-section caption does not state a fixative (datasheet A01197). Record the fixative, fixation duration and tissue processing for each specimen before comparing CYGB staining across cases (standard IHC practice). When an established positive section stains weakly, compare sections with documented processing histories using the same pH 6 citrate retrieval and detection run (datasheet A01197; standard IHC practice). Check whether uneven signal follows tissue edges, folds or poorly preserved areas, which can confound interpretation of chromogenic staining (standard IHC practice). Do not assign a CYGB-specific fixation effect without a controlled comparison (standard IHC practice).
Should CYGB staining be cytoplasmic or nuclear?
Evaluate cytoplasmic staining first: tissue IHC reports general cytoplasmic expression, most abundant in a subset of immune cells (HPA tissue IHC). Nuclear staining is biologically plausible because CYGB is listed in both cytoplasm and nucleus (UniProt Q8WWM9 subcellular location). Supported nuclear-speckle and cytosolic patterns come from cell imaging, so they should guide review of nuclear signal without establishing its appearance in chromogenic tissue sections (HPA subcellular). Compare nuclear and cytoplasmic signal within intact cells, using the counterstain to locate nuclei and excluding folds or damaged areas (standard IHC practice). Record compartments separately when scoring rather than combining them into a single positive call (standard IHC practice).
How can I assess an unexpected CYGB staining pattern without an epitope map?
The supplied record lists a single 1–190 CYGB chain, no isoforms and no transmembrane segment (UniProt Q8WWM9 processing, isoforms and topology). It also lists no glycosylation sites or modified residues, so those annotations do not identify a processing-dependent staining pattern (UniProt Q8WWM9 PTMs). The antibody epitope is not supplied here; avoid assigning a particular stained compartment or cell population to a mapped epitope (datasheet A01197; standard IHC practice). Compare staining with the expected tissue and compartment patterns, then repeat questionable results on adjacent sections with an appropriate negative detection control (HPA tissue IHC; standard IHC practice). Keep retrieval and antibody concentration fixed during that comparison to isolate the source of disagreement (standard IHC practice).
How should I cross-check CYGB IHC findings by immunofluorescence?
Use IF as a separate validation experiment, since the selected antibody evidence here documents chromogenic staining of paraffin sections (datasheet A01197). Multiplex CYGB with a validated marker for the cell population under review; tissue IHC reports prominent staining in a subset of immune cells and high staining in skeletal myocytes (HPA tissue IHC). Choose a fluorophore channel after checking autofluorescence in an unstained section, and include single-label controls when channels may overlap (standard IF practice). CYGB has no transmembrane segment and is reported in cytoplasm and nucleus, so assess permeabilisation for access to intracellular epitopes (UniProt Q8WWM9 topology and subcellular location; standard IF practice). Interpret nuclear speckles against the supported cell-imaging pattern without treating it as proof of identical tissue staining (HPA subcellular).
How do I reduce diffuse or widespread brown CYGB staining?
First compare the stained section with a no-primary control and inspect whether brown deposit follows tissue edges, folds or damaged regions (standard IHC practice). The documented workflow used 10% goat serum blocking, a biotinylated secondary antibody and DAB development (datasheet A01197). Check peroxidase blocking and detection-only controls, then adjust DAB development if background persists across both test and control sections (standard chromogenic IHC practice). Interpret widespread staining cautiously: tissue IHC describes general cytoplasmic expression but greater abundance in a subset of immune cells (HPA tissue IHC). Retain the same retrieval and exposure conditions across comparison sections so changes in background remain interpretable (standard IHC practice).
What is a defensible way to quantify CYGB IHC? ⚠ ANSWER MARKED FOR VERIFICATION
Define the tissue region and cell population before scoring, because reported CYGB staining varies by cell type and compartment (HPA tissue IHC; UniProt Q8WWM9 subcellular location). For identified cells, report the percentage positive and an intensity-weighted H-score on the conventional 0–300 scale, keeping nuclear and cytoplasmic scores separate (standard IHC scoring practice). If the question concerns scattered positive cells, report their density per mm² of viable tissue instead (standard IHC scoring practice). Normalise cell counts to the number of eligible cells and area-based counts to the measured viable area (standard IHC scoring practice). Apply one threshold, counterstain standard and imaging method to all compared sections (standard IHC scoring practice).
When is a CYGB-positive cell convincing rather than an artefact?
A convincing call combines intact-cell staining with the expected cytoplasmic pattern and a plausible cell population; tissue IHC reports general cytoplasmic expression enriched in a subset of immune cells (HPA tissue IHC). High staining in skeletal myocytes and medium staining in colon or duodenal glandular cells provide supplied comparison contexts, whereas listed negative cell populations warrant closer review of a strong signal (HPA tissue IHC). Nuclear signal merits separate assessment because CYGB is also annotated in the nucleus and nuclear speckles are supported in cell imaging (UniProt Q8WWM9 subcellular location; HPA subcellular). Exclude edge deposits and necrotic or folded regions, and use a no-primary control to investigate possible endogenous enzyme signal (standard chromogenic IHC practice). Treat a pattern that follows damaged tissue or appears in the control as unresolved until the staining source is identified (standard IHC practice).
Boster reagents
Best CYGB / Cytoglobin IHC Antibodies
A01197 has IHC images from paraffin sections of mouse intestine and cardiac muscle, rat intestine, and human intestinal cancer tissue (catalog A01197 IHC captions). No IF/ICC data are supplied (catalog A01197 applications).
Real IHC data
Anti-Cytoglobin/CYGB Antibody ®
Cat # A01197
A01197 has IHC images from paraffin sections of mouse intestine and cardiac muscle, rat intestine, and human intestinal cancer tissue (catalog A01197 IHC captions). It is listed for IHC in human, mouse, and rat samples (catalog A01197 applications and reactivity).
Which to pick: Choose A01197 for paraffin-section IHC; its captions document citrate pH 6 retrieval and 1 μg/ml primary antibody, but do not report the fixative (catalog A01197 IHC captions). A01197 is also the cross-species choice listed for human, mouse, and rat; it is a rabbit antibody with no clone specified (catalog A01197 reactivity, host, and clone). No IF/ICC choice is supported by the supplied application list or images (catalog A01197 applications and IF images).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q8WWM9 (CYGB_HUMAN, Cytoglobin).
- Human Protein Atlas. CYGB tissue IHC expression (reliability: Approved).
- Human Protein Atlas. CYGB subcellular location (ICC-IF): Mainly localized to the nuclear speckles. In addition localized to the cytosol..
- Human Protein Atlas. CYGB antibody validation summary (2 antibodies).
- IRP1/ARID3A complex promotes pancreatic cancer chemoresistance by suppressing CYGB-related ferroptosis. Genes & diseases 2026 — PMC13254595.
- Stagnation of histopathological improvement is a predictor of hepatocellular carcinoma development after hepatitis C virus eradication. PloS one 2018 — PMC5849339.
- The expression of cytoglobin as a prognostic factor in gliomas: a retrospective analysis of 88 patients. BMC cancer 2013 — PMC3663650.
- Cytoglobin deficiency potentiates Crb1-mediated retinal degeneration in rd8 mice. Developmental biology 2020 — PMC6995765.
- PubMed PMID:11919282 — UniProt-cited evidence.
- PubMed PMID:12359339 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for CYGB (Cytoglobin). Expected localisation Predominantly cytoplasmic in tissue IHC (HPA tissue IHC). Antigen retrieval: Citrate pH 6 HIER, heat-mediated (datasheet A01197). Validated IHC antibody: A01197 at 0.5-1μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Skeletal muscle (Myocytes); negative Bone marrow. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded mouse intestine tissues; fixative not specified (datasheet A01197); 2. Fixation: Image fixative and duration unreported (datasheet A01197); 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: Citrate pH 6 , 20 min (datasheet A01197); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A01197); 8. Primary antibody: Rabbit anti-CYGB, 0.5-1μg/ml (datasheet A01197); 9. Primary incubation: Overnight at 4 °C (datasheet A01197). Watch for: Fixation: Keep fixation consistent across sections (standard IHC practice; not target-specific); Caveat: Staining and RNA show only medium consistency (HPA tissue IHC); Isoform / epitope: No listed isoforms; one 1–190 chain (UniProt); Regulation: Expression regulation is unreported (UniProt). Evidence: UniProt Q8WWM9, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the nuclear speckles. In addition localized to…, Datasheet A01197, PMC5849339, PMC3663650. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/CYGB/immunohistochemistry-ihc-guide.