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COX6A1 / Cytochrome c oxidase subunit 6A1, mitochondrial · IHC design guide
Design Immunohistochemistry for COX6A1
Plan chromogenic COX6A1 IHC in paraffin sections with the catalog antibody (datasheet A09572-1). Compare high-staining kidney tubules with undetected adipocytes, and check the granular cytoplasmic pattern while accounting for HPA's multi-gene targeting caution (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
COX6A1 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 | Granular cytoplasm (HPA tissue IHC); inner mitochondrial membrane (UniProt) | |
| Staining pattern | Granular cytoplasmic staining in most tissues (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A09572-1) | |
| Positive control | Adrenal gland+4 more · see all | |
| Negative control | Adipose tissue+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent across paraffin sections. (standard IHC practice; not target-specific) | |
| Caveat | Possible multi-gene targeting; verify specificity (HPA tissue IHC) | |
| Regulation | Regulation not annotated (UniProt) | |
| Isoform / epitope | No isoforms; mature chain spans both membrane sides (UniProt) |
Section 1
Recommended COX6A1 IHC & IF Protocols
The catalog antibody’s IHC-P protocol (datasheet A09572-1) is followed by a published COX6A1 IHC protocol for mouse liver sections (PMC12581582).
| Sample | Paraffin-embedded human liver cancer tissue; fixative not specified (datasheet A09572-1) |
| Fixation | Image fixative and duration unreported (datasheet A09572-1); 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 A09572-1); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A09572-1) |
| Primary antibody | Rabbit anti-COX6A1, 1:50 recommended; image 2 μg/ml (datasheet A09572-1) |
| Primary incubation | Overnight at 4 °C (datasheet A09572-1) |
| Detection | HRP-conjugated secondary, DAB chromogen (datasheet A09572-1) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | COX6A1-positive staining in glandular cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression with a granular pattern in most tissues. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA retrieval at pH 8.0 for the catalog antibody (datasheet A09572-1); the published IHC method does not specify retrieval conditions (PMC12581582).
Section 2
What Is the Expected COX6A1 Staining Pattern?
COX6A1 is an inner mitochondrial membrane component with one transmembrane segment at residues 35–59 (UniProt P12074 topology). In paraffin IHC, expect granular cytoplasmic staining in many tissues, including kidney tubular cells and colon glandular cells rated High (HPA tissue IHC). HPA rates the tissue pattern Approved, pending external verification, and cautions that the antibody targets proteins from more than one gene (HPA tissue IHC).
| Granular cytoplasmic staining in kidney tubular or colon glandular cells. | This matches the reported tissue pattern and High staining in those cells (HPA tissue IHC). Assess granules within the expected cells, rather than treating uniform tissue color as equivalent evidence. |
| Predominantly nuclear, membranous, or extracellular deposit. | These compartments conflict with the reported granular cytoplasmic IHC pattern (HPA tissue IHC) and inner mitochondrial membrane location (UniProt P12074 topology). Treat the result as suspect and review controls. |
| Strong staining in adipocytes or skeletal muscle myocytes. | HPA reports these cells as Not detected (HPA tissue IHC). Consider cross-reactivity or endogenous chromogenic activity; the Approved pattern remains pending external verification and carries a multi-gene targeting caution (HPA tissue IHC). |
| Diffuse color across cells and surrounding tissue, with granules hard to distinguish. | This obscures the reported granular cytoplasmic pattern (HPA tissue IHC). Assess whether background also appears in the no-primary control before assigning cellular expression (general IHC practice). |
| No staining in kidney tubular cells or colon glandular cells. | Both are reported High (HPA tissue IHC), so check assay performance before interpreting the sample as negative. An HPA intensity category describes observed staining, not guaranteed detection in every preparation. |
💡Expected COX6A1 appearanceCall a positive result when High, granular cytoplasmic staining is visible in kidney tubular or colon glandular cells (HPA tissue IHC); dominant nuclear color or diffuse tissue-wide deposit is suspect against that pattern (HPA tissue IHC; UniProt P12074 topology).
| Target location and topology | COX6A1 sits in the mitochondrial inner membrane, spanning residues 35–59 (UniProt P12074 topology). Evaluate subcellular pattern as granular cytoplasm in tissue IHC (HPA tissue IHC). |
| Tissue and cell selection | Kidney tubules, colon glands, and caudate neuronal cells are rated High; adipocytes and skeletal muscle myocytes are Not detected (HPA tissue IHC). Select a positive control by cell type, not tissue name alone. |
| Antibody evidence | The reported IHC pattern is Approved but pending external verification; HPA cautions that the antibody targets proteins from more than one gene (HPA tissue IHC). Interpret unexpected staining conservatively. |
| Processed protein | UniProt records a 109-aa precursor and a mature chain at residues 25–109 (UniProt P12074 processing). The payload gives no antibody epitope, so it cannot establish which form is detected. |
| Endogenous chromogenic activity | Endogenous enzyme activity can create apparent IHC color unrelated to the primary antibody (general IHC practice). A no-primary control helps distinguish this from specific granular staining. |
| IF/ICC Q: Where should fluorescence appear? | A: Mainly in mitochondria; HPA additionally reports the principal piece (HPA subcellular ICC-IF). Its supported mitochondrial location carries a caution about antibodies targeting proteins from multiple genes (HPA subcellular ICC-IF). |
| Situation | Likely cause | Next action |
|---|---|---|
| Known-positive kidney tubules show no signal. | Assay detection, retrieval, or antibody conditions may be inadequate (general IHC practice); kidney tubular cells are rated High (HPA tissue IHC). | Check a same-run positive section and detection reagents; review the catalog antibody's IHC-P conditions before interpreting absence (general IHC practice). |
| Granules appear, but nuclear stain dominates. | The dominant compartment disagrees with granular cytoplasmic tissue staining (HPA tissue IHC) and mitochondrial localization (UniProt P12074 topology). | Compare the signal with a no-primary control and a known-positive section; score only a credible cytoplasmic pattern (general IHC practice). |
| Adipocytes stain strongly. | Adipocytes are reported Not detected (HPA tissue IHC); cross-reactivity or endogenous chromogenic activity is possible (general IHC practice). | Check no-primary staining and repeat with appropriate controls before calling the adipocyte signal COX6A1 (general IHC practice). |
| Color spreads across the section without clear cellular granules. | Background obscures the reported granular cytoplasmic pattern (HPA tissue IHC); nonspecific detection is possible (general IHC practice). | Review blocking, washes, antibody concentration, and the no-primary control using standard IHC practice; reassess whether granules remain distinguishable. |
| Only a few cell populations stain within a mixed tissue. | HPA levels are cell specific: lung macrophages are Low, while kidney tubular cells are High (HPA tissue IHC). Tissue-wide scoring can obscure this distinction. | Identify and score the named cell population separately; compare each population with its reported HPA level instead of averaging the section. |
| Two IHC runs give different intensity in the same tissue. | Staining intensity depends on assay conditions (general IHC practice); HPA's Approved pattern is pending external verification (HPA tissue IHC). | Compare the runs' positive and no-primary controls, retrieval, detection, and scoring criteria before attributing the difference to expression (general IHC practice). |
Experiment evidence
IHC & IF images showing COX6A1 staining
A09572-1 · IHCIHC analysis of COX6A1 using anti-COX6A1 antibody (A09572-1). COX6A1 was detected in a paraffin-embedded section of human liver cancer tissu
A09572-1 · IHCIHC analysis of COX6A1 using anti-COX6A1 antibody (A09572-1). COX6A1 was detected in a paraffin-embedded section of human liver tissue. Heat
A09572 · IHCImmunohistochemistry of paraffin-embedded rat cerebellum using COX6A1 antibody at dilution of 1:100 .Perform microwave antigen retrieval wit
A09572 · IHCImmunohistochemistry of paraffin-embedded human liver cancer using COX6A1 antibody at dilution of 1:100 .Perform microwave antigen retrieval
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000111775-COX6A1/tissue/cerebral+cortex#imid_18058231
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000111775-COX6A1/tissue/colon#imid_18058157
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000111775-COX6A1/tissue/liver#imid_18058166
HPA · PancreasHuman Protein Atlas IHC, Pancreas. Source: https://www.proteinatlas.org/ENSG00000111775-COX6A1/tissue/pancreas#imid_18058101
HPA · ICC-IF MCF-7Human Protein Atlas immunofluorescence, MCF-7, antibody HPA062394; green = COX6A1, blue = nucleus, red = microtubules.
HPA · ICC-IF MCF-7Human Protein Atlas immunofluorescence, MCF-7, antibody HPA062394; green = COX6A1, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for COX6A1 IHC & IF
🧪Run colon first and require cytoplasmic staining in its glandular cells (HPA: High in colon glandular cells). Use adipose tissue adipocytes as the negative tissue (HPA: Not detected in adipocytes); on the colon slide, non-glandular stromal cells can serve as an internal background comparator if they show absent or much weaker cytoplasmic signal, but their negative status is not established by the supplied HPA rows.
Positive control tissue: Adrenal gland (Glandular cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show COX6A1 in MCF-7, PC-3, U2OS, Sperm, KOLF2.1J, with annotated localisation: Mitochondria (supported) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only slide; a concentration-matched rabbit IgG isotype control; and, if available, COX6A1 knockout tissue as a biological specificity control (standard IHC practice; selected IHC caption: rabbit primary antibody). For chromogenic colon IHC, block endogenous peroxidase and check whether blood or inflammatory cells contribute brown background before scoring glandular cells (standard IHC practice; selected IHC caption: HRP and DAB detection).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected A09572-1 paraffin-section caption does not report its fixative (selected IHC caption: fixative not stated). The reported IHC procedure used heat retrieval in EDTA at pH 8.0, so assess staining under that retrieval condition; its necessity relative to other conditions is unreported (selected IHC caption: EDTA heat retrieval). The supplied evidence does not establish whether frozen sections or IF/ICC are easier; for colon DAB sections, assess endogenous peroxidase background when interpreting glandular staining (HPA: High in colon glandular cells; standard IHC practice).
HPA tissue IHC evidence for COX6A1
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Pending external verification. Caution, targets protein from more than one gene. Pending external verification.). Rows are taken directly from the HPA tissue chart — click any row's HPA link to view the source.
Positive expression · recommended positive controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adrenal gland | Glandular cells | High | Protein (IHC) | HPA → |
| Appendix | Glandular cells | High | Protein (IHC) | HPA → |
| Caudate | Neuronal cells | High | Protein (IHC) | HPA → |
| Cerebral cortex | Neuropil | High | Protein (IHC) | HPA → |
| Colon | 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 → |
| Ovary | Follicle cells | Not detected | Protein (IHC) | HPA → |
| Placenta | Decidual cells | Not detected | Protein (IHC) | HPA → |
| Skeletal muscle | Myocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced COX6A1 IHC Tips
Troubleshoot COX6A1 staining in paraffin sections by checking retrieval, mitochondrial pattern, cell type, and controls before comparing signal intensity.
How should I adjust retrieval when COX6A1 staining is weak?
Start with heat-mediated retrieval in EDTA at pH 8.0 for this antibody (datasheet A09572-1). The selected paraffin-section image used that condition before staining with 2 μg/ml primary antibody overnight at 4°C (caption A09572-1). If signal is weak, check that heating, cooling, and buffer preparation were consistent across slides before changing retrieval chemistry (standard IHC practice). A different buffer can be tested as a fallback on serial sections, with a known positive section processed alongside each condition (standard IHC practice; HPA: High in kidney tubule cells). Compare granular cytoplasmic signal and tissue preservation together when choosing a condition (HPA: granular cytoplasmic pattern; standard IHC practice).
Can fixation explain faint or uneven COX6A1 staining?
COX6A1-specific sensitivity to fixation is unknown from the supplied evidence; the selected image identifies a paraffin section but does not report its fixative (caption A09572-1). Record the fixative, fixation duration, processing history, and section age for each specimen before attributing weak staining to the target (standard IHC practice). Compare sections prepared under the same conditions, and include a known positive tissue with each run (standard IHC practice; HPA: High in kidney tubule cells). If one batch stains poorly, review morphology and retrieval consistency before altering antibody concentration (standard IHC practice). The caption documents 2 μg/ml antibody after EDTA retrieval at pH 8.0, without establishing a preferred fixative (caption A09572-1).
What staining pattern should count as plausible COX6A1 localisation?
Expect a granular cytoplasmic pattern in tissue sections, consistent with COX6A1 residing in the mitochondrial inner membrane (HPA: granular cytoplasmic expression; UniProt P12074 subcellular location). Assess whether granules lie within cells rather than over nuclei, lumina, or section debris (HPA: granular cytoplasmic expression; standard IHC practice). HPA reports high staining in kidney tubule cells and colon glandular cells, which can help assess the pattern on control sections (HPA: High in kidney tubule cells and colon glandular cells). Diffuse nuclear staining or uniform extracellular deposit warrants review of background and detection controls (UniProt P12074 subcellular location; standard IHC practice). Light counterstaining can help distinguish cytoplasmic granules from nearby tissue structures (standard IHC practice).
Could epitope position or a related subunit affect staining?
COX6A1 has a transmembrane segment at residues 35–59, with residues 25–34 on the matrix side and 60–109 on the intermembrane-space side (UniProt P12074 topology). The supplied record identifies no isoforms or glycosylation sites, and it does not map this antibody’s epitope (UniProt P12074 record; caption A09572-1). Retrieval response therefore cannot identify which side of the membrane the antibody recognizes (UniProt P12074 topology; standard IHC practice). The complex can contain COX6A1 or COX6A2, so assess antibody specificity before assigning a signal uniquely to COX6A1 (UniProt P12074 subunit annotation). HPA also cautions that its tissue antibody targets proteins from more than one gene (HPA: reliability description).
How can IF help examine a questionable IHC pattern?
For a separate IF experiment, pair COX6A1 with a marker identifying the expected cell type, such as a tubule-cell marker when examining kidney sections (HPA: High in kidney tubule cells; standard IF practice). Select fluorophores after checking tissue autofluorescence in unstained sections, and use single-stain controls to assess channel overlap (standard IF practice). COX6A1 spans the inner mitochondrial membrane, with matrix-facing residues 25–34 and intermembrane-facing residues 60–109 (UniProt P12074 topology). Because this antibody’s epitope is unmapped, test permeabilisation conditions that allow access to the relevant mitochondrial membrane face while preserving mitochondrial structure (UniProt P12074 topology; standard IF practice). Assess whether the resulting puncta align with mitochondrial staining before using IF to support the chromogenic interpretation (HPA: mitochondrial location; standard IF practice).
How do I reduce brown haze without losing mitochondrial granules?
First compare the stained section with a no-primary control to identify signal produced by the detection system or tissue (standard IHC practice). The selected procedure used 10% goat serum, a peroxidase-conjugated secondary antibody, and DAB development (caption A09572-1). Apply a peroxidase block where appropriate and keep DAB development consistent across test and control sections (standard chromogenic IHC practice). If haze persists, reassess blocking, washing, and primary-antibody concentration while retaining a positive control to monitor specific granular staining (standard IHC practice; HPA: granular cytoplasmic pattern). The caption’s 2 μg/ml concentration is a tested starting point for this paraffin-section example, not proof that every specimen requires it (caption A09572-1).
What should I measure when comparing COX6A1 across sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and tissue area before scoring, since HPA reports different staining levels across cell types (HPA: tissue IHC profile). An H-score can combine the percentage of cells in each intensity category with its intensity; record the scoring rules before reviewing groups (standard IHC quantification practice). For a simpler readout, report the percentage of positive cells or the density of positive cells per mm² within the same annotated compartment (standard IHC quantification practice). Normalize counts to eligible cells or evaluable tissue area, and exclude folds, necrosis, and tissue loss consistently (standard IHC quantification practice). Keep retrieval, DAB development, and imaging settings comparable across sections, using the granular cytoplasmic pattern to define positivity (HPA: granular cytoplasmic pattern; standard IHC practice).
How can I distinguish true COX6A1 signal from artefact?
A plausible positive shows granular cytoplasmic staining in intact cells, consistent with mitochondrial inner-membrane localisation (HPA: granular cytoplasmic pattern; UniProt P12074 subcellular location). Check a known positive compartment, such as kidney tubule cells, alongside the study section (HPA: High in kidney tubule cells; standard IHC practice). Treat nuclear-only staining, extracellular deposits, and signal confined to section edges or necrotic areas as reasons to inspect the slide and controls (UniProt P12074 subcellular location; standard IHC practice). A no-primary control helps reveal endogenous enzyme activity or detection-system signal during chromogenic development (standard IHC practice). Interpret unexpected cell-type staining cautiously because the HPA tissue result remains pending external verification and carries a multiple-gene targeting caution (HPA: reliability description).
Boster reagents
Best COX6A1 / Cytochrome c oxidase subunit 6A1, mitochondrial IHC Antibodies
Both rabbit polyclonal antibodies have paraffin-section IHC images and list IF/ICC use across human, mouse, and rat; the payload provides no IF images (catalog applications, reactivity, and IHC captions).
Real IHC data
Anti-COX6A1 Antibody
Cat # A09572-1
Real IHC data
Anti-COX6A1 Antibody
Cat # A09572
A09572-1 lists IHC and IF/ICC for human, mouse, and rat, with paraffin-section IHC images from human liver cancer and liver (catalog applications, reactivity, and IHC captions). A09572 lists the same applications and species, with paraffin-section IHC images from rat cerebellum, human liver cancer, and mouse liver (catalog applications, reactivity, and IHC captions).
Which to pick: For human tissue IHC, choose A09572-1 when its documented EDTA pH 8.0 retrieval and 2 μg/ml primary incubation suit your assay; its caption shows paraffin sections but does not report the fixative (A09572-1 IHC captions). For IF/ICC, either rabbit polyclonal antibody is listed: A09572 offers a 1:50–1:200 IF range, while A09572-1 lists 1:50; neither has an IF image in the payload (catalog applications, dilutions, and IF image alts). For IHC across the listed species, choose A09572 because its paraffin-section images cover human, mouse, and rat; its captions do not report the fixative (A09572 IHC captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P12074 (CX6A1_HUMAN, Cytochrome c oxidase subunit 6A1, mitochondrial).
- Human Protein Atlas. COX6A1 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. COX6A1 subcellular location (ICC-IF): Mainly localized to the mitochondria. In addition localized to the principal piece. Caution: Based on antibodies targeting proteins from multiple genes..
- Human Protein Atlas. COX6A1 antibody validation summary (1 antibodies).
- A novel palmitoylation-based molecular signature reveals COX6A1 as a key regulator in metabolic dysfunction-associated steatotic liver disease. Journal of translational medicine 2025 — PMC12581582.
- PLK1 (polo like kinase 1)-dependent autophagy facilitates gefitinib-induced hepatotoxicity by degrading COX6A1 (cytochrome c oxidase subunit 6A1). Autophagy 2021 — PMC8526032.
- PubMed PMID:14702039 — UniProt-cited evidence.
- PubMed PMID:16541075 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for COX6A1 (Cytochrome c oxidase subunit 6A1, mitochondrial). Expected localisation Granular cytoplasm (HPA tissue IHC); inner mitochondrial membrane (UniProt). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A09572-1). Validated IHC antibody: A09572-1 at 1:50 recommended; image 2 μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Glandular cells); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human liver cancer tissue; fixative not specified (datasheet A09572-1); 2. Fixation: Image fixative and duration unreported (datasheet A09572-1); 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 A09572-1); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A09572-1); 8. Primary antibody: Rabbit anti-COX6A1, 1:50 recommended; image 2 μg/ml (datasheet A09572-1); 9. Primary incubation: Overnight at 4 °C (datasheet A09572-1). Watch for: Fixation: Keep fixation consistent across paraffin sections. (standard IHC practice; not target-specific); Caveat: Possible multi-gene targeting; verify specificity (HPA tissue IHC); Isoform / epitope: No isoforms; mature chain spans both membrane sides (UniProt); Regulation: Regulation not annotated (UniProt). Evidence: UniProt P12074, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the mitochondria. In addition localized to the…, Datasheet A09572-1, PMC12581582. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/COX6A1/immunohistochemistry-ihc-guide.