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CCDC69 / Coiled-coil domain-containing protein 69 · IHC design guide
Design Immunohistochemistry for CCDC69
Plan CCDC69 chromogenic IHC on paraffin sections using the IHC-validated antibody at 1–5 μg/mL (datasheet). Compare cytoplasmic staining across cell types while accounting for the uncertain tissue IHC reliability (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
CCDC69 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 staining in most cell types (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic staining across most cell types (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Adrenal gland+4 more · see all | |
| Negative control | Adipose tissue |
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 has low consistency with RNA expression (HPA tissue IHC) | |
| Regulation | Staining-linked regulation unreported (UniProt) | |
| Isoform / epitope | No isoforms listed; chain spans aa 2–296 (UniProt) |
Section 1
Recommended CCDC69 IHC & IF Protocols
The catalog antibody’s IHC-P protocol is followed by three published CCDC69 IHC protocols (PMC7922363; PMC11992169; PMC10571395).
| Sample | Tissue sections; selected-image fixative not specified (standard IHC workflow) |
| Fixation | Image fixative and duration unreported (datasheet A17261); 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 | Chicken anti-CCDC69, 1-5 μg/mL (datasheet A17261) |
| 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 | CCDC69-positive staining in glandular cells of adrenal gland (HPA tissue IHC: Medium). HPA tissue profile: Cytoplasmic expression in most cell types. No signal in the no-primary control. |
💡Decision noteStart with citrate pH 6.0 retrieval at 95–98 °C for 20 min (page retrieval rule); EDTA pH 9.0 is a published alternative (PMC7922363).
Section 2
What Is the Expected CCDC69 Staining Pattern?
CCDC69 is described as cytoplasmic and associated with the spindle and midbody during cell division (UniProt A6NI79). In tissue IHC, expect predominantly cytoplasmic staining in cell populations reported as positive, including glandular and respiratory epithelial cells (HPA tissue IHC). CCDC69 has no annotated transmembrane segment (UniProt A6NI79 topology). Treat the tissue pattern as provisional: HPA rates its IHC reliability Uncertain because staining and RNA expression have low consistency (HPA tissue IHC).
| Cytoplasmic staining in appendix glandular cells or bronchus respiratory epithelial cells. | These cell types have Medium staining in the HPA tissue survey (HPA tissue IHC). Score the compartment and identified cell population together; a positive result remains provisional because the overall tissue IHC profile is rated Uncertain (HPA tissue IHC). |
| A predominantly nuclear signal, with little cytoplasmic staining. | This differs from the reported cytoplasmic tissue pattern (HPA tissue IHC) and the spindle and midbody localisation described during division (UniProt A6NI79). Check staining controls and tissue morphology before treating the nuclear signal as CCDC69; an off-target or detection artefact is possible. |
| Strong staining confined to adipocytes while expected glandular cells are unstained. | HPA reports adipocytes as Not detected and several glandular populations as Medium (HPA tissue IHC). Review cell identification, cross-reactivity and endogenous detection activity; the HPA reliability rating prevents a single discrepant specimen from proving that either signal is false (HPA tissue IHC). |
| Diffuse colour across cells, stroma and tissue edges, without a discernible cellular pattern. | This is difficult to reconcile with the reported cytoplasmic cellular staining (HPA tissue IHC). Assess background in controls and review blocking, washes and chromogen development as general IHC workflow checks; diffuse colour alone does not establish CCDC69 expression. |
| No discernible signal in appendix glandular cells used as a reference specimen. | Those cells showed Medium staining in the HPA survey, so absence warrants a run-level check (HPA tissue IHC). Because the HPA tissue profile is Uncertain, a negative section cannot by itself establish assay failure or genuine absence; inspect tissue preservation and run controls. |
💡Expected CCDC69 appearanceCall a section provisionally positive when identified cells show cytoplasmic staining, such as Medium staining in appendix glandular cells (HPA tissue IHC); isolated nuclear colour or staining confined to adipocytes merits review against the reported pattern (HPA tissue IHC).
| Strength of the tissue reference | HPA rates tissue IHC Uncertain for low agreement between staining and RNA, and lists HPA043648 IHC as Uncertain (HPA tissue IHC; HPA antibodies). Use its cell patterns as comparison points, not definitive expression controls. |
| Cell population and tissue context | HPA reports Medium staining in appendix glandular cells and bronchus respiratory epithelial cells, but Not detected in adipocytes (HPA tissue IHC). Identify the stained cell population before comparing intensities across sections. |
| Compartment and division stage | UniProt places CCDC69 on overlapping interpolar microtubules in early anaphase, at the spindle midzone later, and at the midbody in telophase (UniProt A6NI79). Such structures apply to dividing cells; their absence from a routine section does not negate cytoplasmic staining. |
| IF/ICC comparison: should membrane staining be expected in IHC? | HPA ICC-IF reports plasma membrane as the main location, with Golgi and actin filament staining also approved (HPA subcellular). Its tissue IHC summary instead reports cytoplasmic staining (HPA tissue IHC). Interpret each application against its own evidence; this row does not prescribe an IF protocol. |
| Situation | Likely cause | Next action |
|---|---|---|
| The reference section has no visible cytoplasmic signal. | Appendix glandular cells had Medium staining in HPA, although tissue IHC reliability is Uncertain (HPA tissue IHC). A weak run or a genuinely different specimen remain possible. | Check a processed positive reference and detection controls, then review retrieval, antibody dilution and development using the assay's documented conditions (general IHC practice). Do not infer CCDC69-specific fixation sensitivity. |
| Most of the section shows diffuse chromogen. | The result obscures the reported cellular cytoplasmic pattern (HPA tissue IHC). In general IHC practice, insufficient blocking, washing or overdevelopment can raise background. | Compare negative and reagent controls; optimise blocking, washes and development time as general IHC steps. Reassess cell-restricted staining only after background is controlled. |
| Signal appears mainly in adipocytes. | HPA lists adipocytes as Not detected (HPA tissue IHC). Cell misidentification, cross-reactivity or endogenous detection activity could account for a discrepant result. | Confirm the stained cell type on the counterstained section and inspect detection controls (general IHC practice). Compare with an HPA-reported positive population, while retaining the Uncertain reliability caveat (HPA tissue IHC). |
| The dominant signal is nuclear. | A nuclear-dominant pattern conflicts with HPA's cytoplasmic tissue summary (HPA tissue IHC); UniProt describes spindle and midbody localisation during division (UniProt A6NI79). | Inspect morphology and controls, then reassess localisation in a second HPA-reported cell population (HPA tissue IHC). Treat persistent nuclear-only staining as unresolved rather than scoring it as confirmed CCDC69. |
| A few dividing cells show concentrated central staining. | CCDC69 is described at the spindle midzone in late anaphase and the midbody in telophase (UniProt A6NI79). A concentrated signal can therefore depend on division stage. | Check whether stained structures align with identifiable dividing cells (general histology practice). Record this separately from the predominant cytoplasmic tissue pattern (HPA tissue IHC). |
| An IF image suggests membrane staining, but the IHC section looks cytoplasmic. | HPA reports plasma membrane as the main ICC-IF location, while its tissue IHC summary is cytoplasmic (HPA subcellular; HPA tissue IHC). The observations come from different applications. | Score the paraffin IHC section against the tissue IHC pattern and its Uncertain reliability rating (HPA tissue IHC). Use the IF finding as an application-specific comparison, not a reason to reclassify IHC staining. |
Experiment evidence
IHC & IF images showing CCDC69 staining
A17261 · IHCImmunohistochemistry of CCDC69 in human lung tissue with CCDC69 antibody at 5 μg/mL.
A17261 · IFImmunofluorescence of CCDC69 in human lung tissue with CCDC69 antibody at 20 μg/mL.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000198624-CCDC69/tissue/cerebral+cortex#imid_11871663
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000198624-CCDC69/tissue/colon#imid_11871581
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000198624-CCDC69/tissue/liver#imid_11871590
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000198624-CCDC69/tissue/kidney#imid_11871599
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA052896; green = CCDC69, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA052896; green = CCDC69, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for CCDC69 IHC & IF
🧪Run adrenal gland first and expect glandular cells to stain (HPA: adrenal gland glandular cells, Medium). Use adipose tissue as the negative tissue, with adipocytes showing no detectable staining (HPA: adipose tissue adipocytes, Not detected); on the adrenal slide, cells outside the glandular compartment should show only background if used as internal negative comparators, but their CCDC69 status requires validation (HPA: adrenal gland glandular cells, Medium; standard IHC practice).
Positive control tissue: Adrenal gland (Glandular cells, HPA Medium)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show CCDC69 in A-431, U-251MG, U2OS, with annotated localisation: Plasma membrane (approved) (HPA subcellular).
Technical controls: Include a no-primary (secondary-only) control, an isotype control matched to the primary antibody’s host species and antibody class, and a CCDC69 knockout specimen or validated peptide-block control (standard IHC practice). For adrenal chromogenic IHC, quench endogenous peroxidase and block endogenous biotin if using avidin–biotin detection (standard IHC practice).
⚠️Feasibility: A CCDC69-specific fixation window and retrieval dependence are unreported in the supplied evidence; optimise antigen retrieval empirically for paraffin IHC (supplied evidence; standard IHC practice). The selected A17261 lung tissue-IHC caption does not report a fixative (selected A17261 caption); HPA has ICC-IF images from A-431, U-251MG and U2OS, but the supplied evidence does not establish whether frozen sections or IF are easier (HPA: subcellular). Endogenous brown pigment in adrenal tissue can resemble chromogenic staining, so compare it with the no-primary control (standard IHC practice).
HPA tissue IHC evidence for CCDC69
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Uncertain — 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 |
|---|---|---|---|---|
| Adrenal gland | Glandular cells | Medium | Protein (IHC) | HPA → |
| Appendix | Glandular cells | Medium | Protein (IHC) | HPA → |
| Breast | Glandular cells | Medium | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | Medium | Protein (IHC) | HPA → |
| Caudate | Glial cells | Medium | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced CCDC69 IHC Tips
Troubleshoot CCDC69 chromogenic IHC by checking retrieval, compartment, and controls before comparing staining across sections.
How should I retrieve CCDC69 in paraffin sections when staining is weak?
Start with citrate pH 6.0 heat-induced retrieval at 95–98 °C for 20 min (page retrieval rule: cytoplasmic / membrane antigen). Let sections cool in the retrieval solution, then keep the heating and cooling schedule identical across comparison slides (standard IHC practice). The selected lung tissue caption reports staining at 5 µg/mL but does not report retrieval conditions, so that image cannot establish an optimal retrieval method (caption: A17261). If staining remains weak, compare a second retrieval condition on matched sections while holding antibody concentration and detection constant, and reject conditions that damage morphology or raise background (standard IHC practice).
Could fixation explain variable CCDC69 staining between paraffin blocks?
Target-specific fixation sensitivity is unknown because no CCDC69 fixation experiment is supplied (evidence payload: fixation unreported). The selected lung tissue caption names a tissue and antibody concentration of 5 µg/mL, but states no fixative, so it cannot resolve fixation effects (caption: A17261). Compare sections with documented processing histories using the same citrate pH 6.0 retrieval and detection run, then assess morphology alongside staining (page retrieval rule; standard IHC practice). Include a reference section in each run and record fixation and processing details before attributing intensity differences to CCDC69 abundance (standard IHC practice).
Which staining compartment should I expect for CCDC69 in tissue IHC?
Assess cytoplasmic staining first: tissue IHC describes cytoplasmic expression in most cell types, although its reliability is uncertain (HPA tissue IHC). Spindle and midbody staining may appear in appropriately staged dividing cells because CCDC69 localizes to overlapping interpolar microtubules in early anaphase and concentrates at the midbody in telophase (UniProt A6NI79 localisation). HPA cell imaging also reports a mainly plasma membrane location, with Golgi and actin filament signals, creating a compartment discrepancy that tissue IHC alone cannot settle (HPA subcellular). Record cytoplasmic, membrane, and mitotic patterns separately, and require morphology and controls before interpreting a compartment-restricted signal (standard IHC practice).
How can epitope uncertainty affect CCDC69 IHC interpretation?
The record lists a 296-amino-acid CCDC69 protein and no annotated isoforms, but supplies no antibody epitope, so staining cannot be assigned to a particular region (UniProt A6NI79 record; antibody payload). The protein has no annotated transmembrane segment, and phosphoserines are reported at residues 154 and 241 (UniProt A6NI79 topology and modified residues). Without epitope mapping, do not infer whether fixation or retrieval preserves a phosphorylated or unphosphorylated form (antibody payload; standard IHC interpretation). If specificity is uncertain, compare independently validated reagents where available and interpret agreement by cell type and compartment rather than intensity alone (standard IHC practice).
How should I check a CCDC69 IHC pattern by multiplex IF?
Pair CCDC69 with a marker for the expected cell type, such as a respiratory epithelial marker when examining bronchus, where respiratory epithelial staining is reported at medium level (HPA tissue IHC). Choose a far-red fluorophore for a weak channel and inspect an unstained section to gauge tissue autofluorescence before setting exposure (standard IF practice). Use controlled permeabilisation when testing intracellular cytoplasmic or spindle staining, since CCDC69 has no transmembrane segment and UniProt places it at the spindle and midbody (UniProt A6NI79 topology and localisation; standard IF practice). Evaluate any membrane-like signal separately because HPA cell imaging reports plasma membrane localisation, while the tissue IHC profile is mainly cytoplasmic and uncertain (HPA subcellular; HPA tissue IHC).
What should I check when CCDC69 DAB staining looks diffuse?
Run a no-primary control and check whether color persists in the same structures; residual signal can reflect detection background rather than CCDC69 (standard IHC practice). Apply a peroxidase block before chromogenic detection and develop DAB for the same interval across matched sections, while monitoring for excessive color deposition (standard IHC practice). Compare the suspect section with a reference section processed in the same run, and inspect tissue edges and damaged areas separately (standard IHC practice). The selected lung caption reports 5 µg/mL for A17261, but provides no blocking or DAB conditions to treat as target-specific settings (caption: A17261).
How should I score CCDC69 staining across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, because HPA describes mainly cytoplasmic tissue staining but also flags low agreement with RNA data (HPA tissue IHC). For cytoplasmic chromogenic signal, report the percentage of positive cells and an H-score derived from intensity categories; count spindle or midbody-positive cells separately when morphology permits (standard IHC quantification; UniProt A6NI79 localisation). Normalize counts to the number of evaluable cells in the same annotated compartment, or express discrete positive structures per mm² of viable tissue (standard IHC quantification). Keep acquisition, DAB development, section inclusion rules, and thresholds consistent across groups, and report how uncertain or damaged areas were excluded (standard IHC practice).
When is a CCDC69-positive IHC result convincing?
A convincing result follows a reproducible cellular pattern with clean controls: cytoplasmic staining is reported in most cell types, but HPA rates the tissue IHC pattern uncertain (HPA tissue IHC; standard IHC practice). Spindle-midzone or midbody staining should align with recognizable anaphase or telophase figures, matching the recorded mitotic localisation (UniProt A6NI79 localisation). Treat strong nuclear-only color, isolated tissue-edge staining, necrotic areas, or signal reproduced in a no-primary control as reasons to investigate artefact (standard IHC practice; UniProt A6NI79 localisation). Also check peroxidase-block controls before attributing chromogen deposition to CCDC69, and avoid using staining intensity alone as proof of abundance (standard IHC practice).
Boster reagents
Best CCDC69 / Coiled-coil domain-containing protein 69 IHC Antibodies
A17261 has IHC and IF images from human lung tissue (catalog image captions). Human, mouse, and rat reactivity is listed, but tissue imaging evidence is human only (catalog reactivity and image captions).
Real IHC data
Anti-CCDC69 Antibody
Cat # A17261
A17261 is listed for IHC-P and IF, with human, mouse, and rat reactivity (catalog applications and reactivity). Its images show human lung IHC at 5 μg/mL and IF at 20 μg/mL (A17261 image captions).
Which to pick: Choose A17261 for paraffin-section IHC because IHC-P is listed and its own IHC image shows staining in human lung tissue (catalog applications; A17261 IHC image caption); the fixative is unreported (A17261 IHC image caption). Choose A17261 for tissue IF because its own IF image shows human lung staining; ICC validation is unreported (A17261 IF image caption; catalog applications). For mouse or rat work, A17261 lists reactivity in both species, but the supplied IHC and IF images show human tissue only (catalog reactivity; A17261 image captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry A6NI79 (CCD69_HUMAN, Coiled-coil domain-containing protein 69).
- Human Protein Atlas. CCDC69 tissue IHC expression (reliability: Uncertain).
- Human Protein Atlas. CCDC69 subcellular location (ICC-IF): Mainly localized to the plasma membrane. In addition localized to the Golgi apparatus and actin filaments..
- Human Protein Atlas. CCDC69 antibody validation summary (2 antibodies).
- Study on CCDC69 interfering with the prognosis of patients with breast cancer through PPAR signal pathway. European journal of histochemistry : EJH 2021 — PMC7922363.
- Identification of M1 macrophage infiltration-related genes for immunotherapy in Her2-positive breast cancer based on bioinformatics analysis and machine learning. Scientific reports 2025 — PMC11992169.
- High expression of CCDC69 is correlated with immunotherapy response and protective effects on breast cancer. BMC cancer 2023 — PMC10571395.
- PubMed PMID:14702039 — UniProt-cited evidence.
- PubMed PMID:15372022 — UniProt-cited evidence.
- PubMed PMID:20962590 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for CCDC69 (Coiled-coil domain-containing protein 69). Expected localisation Cytoplasmic staining in most cell types (HPA tissue IHC). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: A17261 at 1-5 μg/mL (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Glandular cells); negative Adipose tissue. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Tissue sections; selected-image fixative not specified (standard IHC workflow); 2. Fixation: Image fixative and duration unreported (datasheet A17261); 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: Chicken anti-CCDC69, 1-5 μg/mL (datasheet A17261); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across sections. (standard IHC practice; not target-specific); Caveat: Staining has low consistency with RNA expression (HPA tissue IHC); Isoform / epitope: No isoforms listed; chain spans aa 2–296 (UniProt); Regulation: Staining-linked regulation unreported (UniProt). Evidence: UniProt A6NI79, HPA tissue IHC · Uncertain, HPA ICC-IF · Mainly localized to the plasma membrane. In addition localized to…, Datasheet A17261, PMC7922363, PMC11992169. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/CCDC69/immunohistochemistry-ihc-guide.