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ANKRD26 / Ankyrin repeat domain-containing protein 26 · IHC design guide
Design Immunohistochemistry for ANKRD26
Plan ANKRD26 staining in paraffin sections using the cytoplasmic, often granular tissue pattern (HPA tissue IHC). Compare positive and negative tissues from the tissue profile, and titrate the IHC-validated antibody at 1:100–1:300 (HPA tissue IHC; datasheet).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
ANKRD26 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, often granular (HPA tissue IHC) | |
| Staining pattern | Glandular and neuronal cytoplasm, often granular (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Appendix+4 more · see all | |
| Negative control | Adipose tissue+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A09289-1) | |
| Caveat | Staining and RNA expression show medium consistency (HPA tissue IHC) | |
| Regulation | Expression regulation not specified (UniProt) | |
| Isoform / epitope | 2 isoforms; epitope coverage unspecified (UniProt) |
Section 1
Recommended ANKRD26 IHC & IF Protocols
The catalog antibody’s IHC-P protocol is accompanied by published ANKRD26 staining protocols for zebrafish paraffin sections and mouse brain cryosections (PMC12067082; PMC4601608).
| Sample | Paraffin-embedded human brain tissue; fixative not specified (datasheet A09289-1) |
| Fixation | Image fixative and duration unreported (datasheet A09289-1); 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 citrate buffer, pH 6.0, 20 min at 95–98 °C (standard rule: cytoplasmic / membrane 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 anti-ANKRD26, 1:100-1:300 (datasheet A09289-1) |
| 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 | ANKRD26-positive staining in glandular cells of appendix (HPA tissue IHC: High). HPA tissue profile: General cytoplasmic expression, which often had a granular pattern. No signal in the no-primary control. |
💡Decision noteFor paraffin sections, start with citrate pH 6.0 retrieval at 95–98 °C for 20 min (page antigen-retrieval rule).
Section 2
What Is the Expected ANKRD26 Staining Pattern?
In paraffin section IHC, expect mainly cytoplasmic ANKRD26 staining, often granular, with high staining in several glandular cell populations and cerebral cortex neurons (HPA tissue IHC). ICC-IF places ANKRD26 mainly at the Golgi apparatus, with additional vesicular and cytosolic signal (HPA subcellular ICC-IF). UniProt annotates no transmembrane segment or subcellular location (UniProt Q9UPS8 topology). HPA rates tissue IHC “Approved,” with medium staining–RNA consistency and external verification pending (HPA tissue IHC).
| Granular cytoplasmic staining in colon or duodenal glandular cells, or cerebral cortex neurons. | This matches the reported IHC compartment and high-staining cell populations (HPA tissue IHC). Judge the distribution within the named cells; neighbouring cell populations need their own reference. |
| Predominantly nuclear or sharply surface-membrane staining, with little cytoplasmic signal. | This differs from the general cytoplasmic IHC profile (HPA tissue IHC) and the Golgi, vesicular and cytosolic ICC-IF locations (HPA subcellular ICC-IF). Check controls before treating it as ANKRD26. |
| Strong staining in adipocytes, bone marrow hematopoietic cells or liver cholangiocytes. | Those specific cell populations were reported as not detected (HPA tissue IHC). Unexpected staining may reflect cross-reactivity or endogenous detection activity (general IHC practice); it does not establish that every cell in those tissues is negative. |
| Diffuse colour across cells, extracellular spaces and tissue edges. | A field-wide deposit is difficult to reconcile with the reported cell-associated cytoplasmic pattern (HPA tissue IHC). Background from detection reagents or insufficient blocking is possible (general IHC practice). |
| No staining in colon glandular cells on an otherwise interpretable section. | Colon glandular cells were scored high (HPA tissue IHC). Review tissue preservation, retrieval and detection controls before concluding that the sample lacks ANKRD26 (general IHC practice); an HPA level is not a guarantee for every specimen. |
💡Expected ANKRD26 appearanceA convincing positive shows predominantly cytoplasmic, often granular staining in high-scoring glandular cells or cerebral cortex neurons (HPA tissue IHC); dominant nuclear, surface-membrane or field-wide colour warrants a control check (HPA tissue IHC; general IHC practice).
| Cell-type reference | High staining is reported in appendix, colon, duodenum, gallbladder, rectum, salivary gland and seminal vesicle glandular cells, plus cerebral cortex neurons (HPA tissue IHC). Score the specified cells rather than whole tissues. |
| Lower and absent reference signals | Thyroid and adrenal glandular cells, lung alveolar cells, testis seminiferous-duct cells and soft-tissue fibroblasts scored low; selected adipocytes, hematopoietic cells, cholangiocytes, ovarian stromal cells and splenic red-pulp cells were not detected (HPA tissue IHC). |
| IHC evidence strength | The tissue profile is “Approved,” with medium staining–RNA consistency and external verification pending (HPA tissue IHC). Use positive and negative cell-type controls when interpreting a new specimen (general IHC practice). |
| IF/ICC interpretation | Does IF/ICC have the same readout? ICC-IF shows mainly Golgi signal, with additional vesicular and cytosolic signal (HPA subcellular ICC-IF). That cellular localisation helps interpret IF/ICC but does not prescribe an IHC-P staining pattern or an IF protocol. |
| Topology and isoforms | ANKRD26 has no annotated transmembrane segment or signal peptide and has two annotated isoforms (UniProt Q9UPS8). The record supplies no antibody epitope or isoform-specific IHC pattern, so neither isoform can be assigned from staining alone. |
| Antibody validation | HPA040654 and HPA041460 are each IHC “Approved”; only HPA040654 is also ICC “Approved” in the supplied antibody record (HPA antibodies). Approval supports use of those readouts but does not establish target-specific fixation sensitivity. |
| Situation | Likely cause | Next action |
|---|---|---|
| High-scoring glandular cells show no chromogenic signal. | A failed staining run or an unsuitable retrieval or detection condition is possible (general IHC practice); those cells score high in several tissues (HPA tissue IHC). | Run a high-scoring reference section alongside the sample and verify the primary, detection reagents, retrieval step and counterstain (HPA tissue IHC; general IHC practice). |
| Most nuclei stain while cytoplasm stays pale. | The distribution conflicts with the general cytoplasmic tissue profile (HPA tissue IHC); nonspecific staining or interpretation of counterstain is possible (general IHC practice). | Compare a section processed without primary antibody, inspect the chromogen separately from the counterstain, and reassess cytoplasmic signal (general IHC practice). |
| Cholangiocytes or adipocytes stain strongly. | Those cells were reported as not detected (HPA tissue IHC); cross-reactivity or endogenous detection activity may contribute (general IHC practice). | Confirm the cell identity, check a no-primary control and use an appropriate endogenous-activity block for the detection system (general IHC practice). |
| The entire section has granular or diffuse colour. | Uniform deposit obscures the cell-restricted pattern reported by HPA (HPA tissue IHC); inadequate blocking, washing or reagent concentration may raise background (general IHC practice). | Compare no-primary and positive controls; review blocking, wash steps and detection conditions before scoring individual cells (general IHC practice). |
| A weak result in thyroid glandular cells seems negative. | Those cells are reported at low staining, so a faint result can fit the reference (HPA tissue IHC). Background and counterstain can make weak chromogen hard to judge (general IHC practice). | Compare the same run with a high-scoring glandular reference and its no-primary control; score signal above background within the named cells (HPA tissue IHC; general IHC practice). |
| Golgi-like puncta appear in IF/ICC but are hard to resolve in IHC-P. | Golgi is the main ICC-IF location, whereas tissue IHC is described more broadly as often granular and cytoplasmic (HPA subcellular ICC-IF; HPA tissue IHC). | Interpret each application against its own reference: assess cytoplasmic cell-type staining for IHC-P and consult the separate IF/ICC guide for fluorescence interpretation (HPA tissue IHC; HPA subcellular ICC-IF). |
Experiment evidence
IHC & IF images showing ANKRD26 staining
A09289-1 · IHCImmunohistochemistry analysis of paraffin-embedded human brain, using ANKRD26 Antibody. The picture on the right is blocked with the synthes
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000107890-ANKRD26/tissue/cerebral+cortex#imid_10595477
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000107890-ANKRD26/tissue/colon#imid_10595411
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000107890-ANKRD26/tissue/liver#imid_10595404
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000107890-ANKRD26/tissue/kidney#imid_10595405
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA040654; green = ANKRD26, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA040654; green = ANKRD26, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for ANKRD26 IHC & IF
🧪Run colon first: glandular cells should stain strongly (HPA: High in colon glandular cells). Use liver cholangiocytes as the negative tissue (HPA: Not detected in liver cholangiocytes); on the colon slide, compare glandular staining with adjacent non-glandular cells as an internal background reference.
Positive control tissue: Appendix (Glandular cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show ANKRD26 in A-431, U-251MG, U2OS, RPTEC/TERT1, hTERT-RPE1 (serum starved), with annotated localisation: Golgi apparatus (approved) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control and an irrelevant antibody matched to the primary antibody’s host species, immunoglobulin class, and monoclonal or polyclonal format (standard IHC practice). Add a peptide-block control for the catalog antibody (A09289-1 tissue-IHC caption: staining blocked with synthesized peptide), and quench endogenous peroxidase before chromogenic detection in colon sections (standard IHC practice).
⚠️Feasibility: The supplied evidence reports no target-specific fixation window or antigen-retrieval dependency; the paraffin-section brain caption does not report a fixative (A09289-1 tissue-IHC caption). Frozen-section performance is unreported, so there is no basis to call it easier than paraffin IHC; ICC-IF images exist in the listed HPA cell lines, but they do not establish relative ease for tissue sections (HPA subcellular record). In colon, endogenous peroxidase in inflammatory cells can complicate chromogenic scoring (standard IHC practice).
HPA tissue IHC evidence for ANKRD26
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — 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 |
|---|---|---|---|---|
| Appendix | Glandular cells | High | Protein (IHC) | HPA → |
| Cerebral cortex | Neuronal cells | High | Protein (IHC) | HPA → |
| Colon | Glandular cells | High | Protein (IHC) | HPA → |
| Duodenum | Glandular cells | High | Protein (IHC) | HPA → |
| Gallbladder | Glandular cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Liver | Cholangiocytes | Not detected | Protein (IHC) | HPA → |
| Ovary | Ovarian stroma cells | Not detected | Protein (IHC) | HPA → |
| Spleen | Cells in red pulp | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced ANKRD26 IHC Tips
Troubleshoot ANKRD26 staining in paraffin sections using compartment, cell type and control patterns alongside the chromogenic signal.
What retrieval conditions should I try when ANKRD26 staining is weak?
Use citrate buffer at pH 6.0 for heat-induced retrieval at 95–98 °C for 20 min (page retrieval rule). Let sections cool in buffer, then compare them with an otherwise identical section processed without retrieval to judge whether heating improves signal or raises background (standard IHC practice). The selected antibody has a paraffin-section image in human brain, but its caption gives no retrieval conditions, so treat this setting as a starting condition rather than a product-validated method (catalog antibody A09289-1 caption). If staining remains weak, vary heating time on adjacent sections before changing buffer chemistry, and score signal in comparable neuronal cells (HPA: High in cerebral-cortex neuronal cells; standard IHC practice).
How should I troubleshoot ANKRD26 staining across differently fixed specimens?
Target-specific fixation sensitivity is unknown: the selected brain image identifies paraffin embedding but does not report the fixative (catalog antibody A09289-1 caption). Record fixative, fixation duration and processing history for each specimen, and compare matched sections processed with the same citrate pH 6.0 retrieval and detection conditions (page retrieval rule; standard IHC practice). If signal differs, test adjacent sections with a controlled retrieval-time series while keeping antibody dilution and chromogen development constant (standard IHC practice). Do not attribute a fixation effect to ANKRD26 from its phosphorylation sites or from tissue staining patterns; those records do not measure fixation sensitivity (UniProt Q9UPS8 modified residues; HPA tissue IHC).
Which staining compartment is plausible for ANKRD26 in tissue sections?
Look first for cytoplasmic staining, sometimes granular, in the relevant cells (HPA tissue IHC: general cytoplasmic expression, often granular). Golgi-associated and vesicular signal is also plausible because cell imaging places ANKRD26 mainly at the Golgi apparatus, with additional vesicle and cytosol localisation (HPA subcellular: approved locations). In brightfield sections, assess whether granules sit within cell boundaries and recur across comparable cells; a perinuclear accent alone cannot establish Golgi identity at routine IHC resolution (standard IHC interpretation). ANKRD26 has no annotated transmembrane segment, so isolated crisp plasma-membrane staining merits review against controls and morphology before assignment (UniProt Q9UPS8 topology; standard IHC interpretation).
Could epitope choice explain discordant ANKRD26 IHC results?
ANKRD26 has 2 listed isoforms, but the supplied product caption does not locate the antibody epitope or establish isoform coverage (UniProt Q9UPS8 isoforms; catalog antibody A09289-1 caption). Check the antibody's documented immunogen against both isoform sequences before claiming that a stain represents total ANKRD26 (standard antibody validation practice). Several phosphoserines are recorded, including residues 11, 15, 241, 261 and 489, but their effect on this antibody's staining is unreported (UniProt Q9UPS8 modified residues; catalog antibody A09289-1 caption). When results disagree, compare adjacent sections under identical retrieval and detection settings, then seek an independently validated epitope or orthogonal evidence (standard IHC practice).
How can IF help resolve an ambiguous ANKRD26 chromogenic pattern?
On a separately validated IF preparation, multiplex ANKRD26 with a neuronal-cell marker when investigating the cerebral cortex, where neuronal cells stain strongly by tissue IHC (HPA tissue IHC: High in cerebral-cortex neuronal cells; standard IF practice). Choose spectrally separated fluorophores and place the weaker signal in a channel with low measured tissue autofluorescence, using single-stain controls to assess bleed-through (standard IF practice). For an intracellular epitope, include a carefully titrated permeabilisation step after fixation; do not assume that a membrane-side protocol applies, because ANKRD26 lacks a transmembrane segment (UniProt Q9UPS8 topology; standard IF practice). Compare compartment patterns cautiously: approved cell-imaging locations include Golgi, vesicles and cytosol, whereas the paraffin brain caption supplies no IF protocol (HPA subcellular; catalog antibody A09289-1 caption).
How do I reduce diffuse or granular background without losing ANKRD26 signal?
Run no-primary and matched control sections to separate detection-system staining from primary-antibody-dependent signal (standard IHC practice). In a peroxidase-based chromogenic workflow, block endogenous peroxidase before applying the detection reagent and inspect pigment or precipitate independently of cellular staining (standard IHC practice). If diffuse color persists, titrate the primary antibody and detection exposure while keeping citrate pH 6.0 retrieval fixed for the comparison (page retrieval rule; standard IHC practice). Preserve interpretable granular cytoplasmic signal in comparable cells, since that pattern is reported for ANKRD26; do not call every granule background solely because it is punctate (HPA tissue IHC: often granular cytoplasmic expression).
What should I measure when comparing ANKRD26 staining between sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the scored cell population and compartment before analysis; for cerebral cortex, neuronal cells provide a documented high-staining population (HPA tissue IHC: High in cerebral-cortex neuronal cells; standard IHC practice). An H-score can combine the percentages of cells at intensity 0–3, while percent-positive cells or positive-cell density per mm² offers simpler alternatives (standard IHC scoring practice). Normalize each readout to the number of eligible cells or the measured tissue area, and use identical thresholds, retrieval and chromogen development across compared sections (standard IHC practice). Report cytoplasmic and granular signal consistently rather than mixing it with unstained nuclei or unrelated tissue compartments (HPA tissue IHC: cytoplasmic, often granular; standard IHC scoring practice).
When should I doubt an apparently positive ANKRD26 stain?
A credible pattern should be intracellular and broadly compatible with cytoplasmic, sometimes granular tissue staining; cell imaging also supports Golgi, vesicle and cytosol locations (HPA tissue IHC; HPA subcellular). Strong neuronal-cell staining in cerebral cortex is plausible, whereas an isolated signal in a cell population recorded as not detected, such as adipocytes, needs additional validation (HPA tissue IHC: High in cerebral-cortex neuronal cells; Not detected in adipocytes). Question staining restricted to section edges, necrotic regions or no-primary controls, and check endogenous enzyme activity in peroxidase detection (standard IHC interpretation). Peptide blocking was shown for the selected paraffin-brain image, but that control alone does not establish specificity in every specimen or processing condition (catalog antibody A09289-1 caption; standard antibody validation practice).
Boster reagents
Best ANKRD26 / Ankyrin repeat domain-containing protein 26 IHC Antibodies
A09289-1 has an ANKRD26 IHC figure from paraffin-embedded human brain (IHC image caption). IF is listed, with human and mouse reactivity, but no IF figure is supplied (catalog applications/reactivity; IF image alts).
Real IHC data
Anti-ANKRD26 Antibody
Cat # A09289-1
A09289-1 will render with an IHC figure from paraffin-embedded human brain, including a peptide-blocked comparison (IHC image caption). It lists IHC and IF applications and human and mouse reactivity; no IF image is supplied (catalog applications/reactivity; IF image alts).
Which to pick: For tissue IHC, choose A09289-1 based on its own paraffin-section figure; the fixative is unreported (IHC image caption). For IF, A09289-1 lists the application, but ICC and an IF figure are unreported (catalog applications; IF image alts). For cross-species work, A09289-1 lists human and mouse reactivity, though its displayed IHC evidence is human only; it is rabbit-hosted, and clonality is unreported (catalog reactivity/host/clone; IHC image caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q9UPS8 (ANR26_HUMAN, Ankyrin repeat domain-containing protein 26).
- Human Protein Atlas. ANKRD26 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. ANKRD26 subcellular location (ICC-IF): Mainly localized to the Golgi apparatus. In addition localized to vesicles and cytosol..
- Human Protein Atlas. ANKRD26 antibody validation summary (2 antibodies).
- Modeling ANKRD26 5'-UTR mutation-related thrombocytopenia. Disease models & mechanisms 2025 — PMC12067082.
- A novel form of ciliopathy underlies hyperphagia and obesity in Ankrd26 knockout mice. Brain structure & function 2015 — PMC4601608.
- PubMed PMID:10470851 — UniProt-cited evidence.
- PubMed PMID:15164054 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for ANKRD26 (Ankyrin repeat domain-containing protein 26). Expected localisation Cytoplasmic, often granular (HPA tissue IHC). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: A09289-1 at 1:100-1:300 (the product's own IHC validation image is shown). Controls (HPA): positive Appendix (Glandular cells); negative Adipose tissue. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Paraffin-embedded human brain tissue; fixative not specified (datasheet A09289-1); 2. Fixation: Image fixative and duration unreported (datasheet A09289-1); 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-induced epitope retrieval in citrate buffer, pH 6.0 , 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 anti-ANKRD26, 1:100-1:300 (datasheet A09289-1); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A09289-1); Caveat: Staining and RNA expression show medium consistency (HPA tissue IHC); Isoform / epitope: 2 isoforms; epitope coverage unspecified (UniProt); Regulation: Expression regulation not specified (UniProt). Evidence: UniProt Q9UPS8, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the Golgi apparatus. In addition localized to…, Datasheet A09289-1, PMC12067082, PMC4601608. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/ANKRD26/immunohistochemistry-ihc-guide.