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OPA1 / Dynamin-like GTPase OPA1, mitochondrial · IHC design guide
Design Immunohistochemistry for OPA1
Plan OPA1 staining in paraffin sections around the granular cytoplasmic pattern seen in most tissues (HPA tissue IHC). Use the IHC-validated antibody and assess cell-specific signal against the expected tissue pattern (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
OPA1 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) | |
| Staining pattern | Granular cytoplasmic staining in most tissues (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet PB9731) | |
| Positive control | Appendix+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 medium concordance with RNA (HPA tissue IHC) | |
| Regulation | Low tissue specificity (HPA tissue RNA) | |
| Isoform / epitope | Six isoforms; long and cleaved short forms may differ at an unknown epitope (UniProt) |
Section 1
Recommended OPA1 IHC & IF Protocols
The catalog antibody has an IHC-P protocol; the published options below report OPA1 staining in heart tissue (PMC9110818) and tissue sections (PMC5555107).
| Sample | Paraffin-embedded Mouse Liver tissue; fixative not specified (datasheet PB9731) |
| Fixation | Image fixative and duration unreported (datasheet PB9731); 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 PB9731); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet PB9731) |
| Primary antibody | Rabbit anti-OPA1, 0.5-1μg/ml (datasheet PB9731) |
| Primary incubation | Overnight at 4 °C (datasheet PB9731) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet PB9731) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | OPA1-positive staining in glandular cells of appendix (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: PB9731); citrate at pH 6.0 is a published alternative (PMC5555107).
Section 2
What Is the Expected OPA1 Staining Pattern?
OPA1 is an inner mitochondrial membrane protein with a form in the intermembrane space (UniProt O60313 topology and processing). In paraffin section IHC, expect granular cytoplasmic staining in many tissues, including strong staining in heart cardiomyocytes and kidney tubular cells (HPA tissue IHC). HPA rates the tissue pattern Supported, with medium consistency between antibody staining and RNA expression (HPA tissue IHC).
| Granular cytoplasmic signal in cardiomyocytes or kidney tubular cells, with limited surrounding background. | This matches HPA's reported High staining in those cells and its granular cytoplasmic pattern across most tissues (HPA tissue IHC). Granules are consistent with mitochondrial localization, although chromogenic IHC does not resolve the inner membrane or intermembrane space (UniProt O60313 topology; general IHC practice). |
| Predominantly nuclear signal in a paraffin section, with little granular cytoplasmic signal. | This does not match HPA's usual tissue IHC pattern; check whether the signal is specific before scoring it as OPA1 (HPA tissue IHC; general IHC practice). Nuclear staining cannot be dismissed solely by location: HPA reports approved nucleoplasmic localization in ICC-IF (HPA subcellular ICC-IF). |
| Strong staining confined to a cell type reported as negative, such as adipocytes. | HPA reports OPA1 as Not detected in adipocytes (HPA tissue IHC). Review cell identification and controls; cross-reactivity or endogenous chromogenic activity can produce apparent positives (general IHC practice). A single unexpected cell population does not establish a new OPA1 tissue pattern. |
| Uniform diffuse color across cells and extracellular areas, without discernible cytoplasmic granules. | This is difficult to reconcile with the reported granular cytoplasmic tissue pattern (HPA tissue IHC). Assess background from nonspecific antibody binding or detection chemistry before interpreting intensity (general IHC practice); diffuse color alone cannot identify which step is responsible. |
| No signal in a section expected to be strongly positive, such as heart cardiomyocytes or kidney tubular cells. | HPA reports High staining in both cell populations (HPA tissue IHC). Check section quality, antibody and detection controls, and the antigen retrieval conditions used for the assay (general IHC practice). Absence of signal alone does not show that OPA1 is absent. |
💡Expected OPA1 appearanceCall a positive IHC result when known positive cells show distinct granular cytoplasmic staining, especially strong staining in cardiomyocytes or kidney tubules; diffuse deposit or isolated unexpected cell staining needs control review (HPA tissue IHC; general IHC practice).
| Subcellular location | OPA1 has an inner membrane segment and intermembrane space regions (UniProt O60313 topology). At light microscope resolution, interpret the expected tissue IHC signal as cytoplasmic granularity rather than a separately identifiable mitochondrial membrane outline (HPA tissue IHC; general IHC practice). |
| Proteolytic processing | UniProt describes a membrane-tethered long form and a soluble short form in the mitochondrial intermembrane space (UniProt O60313 processing and function). The supplied sources give no antibody epitope, so an IHC stain cannot be assigned to either form from appearance alone. |
| Tissue and cell selection | HPA reports High staining in several glandular cell populations, cardiomyocytes, kidney tubular cells and ovarian follicle cells; adipocytes are Not detected (HPA tissue IHC). HPA also reports low tissue RNA specificity, so choose comparison cells by their reported protein staining level (HPA tissue IHC). |
| Assay-specific location evidence | HPA describes granular cytoplasmic tissue IHC, while its ICC-IF summary lists nucleoplasm as approved and mitochondria as supported (HPA tissue IHC; HPA subcellular ICC-IF). Interpret each compartment claim in its assay context; ICC-IF localization does not establish the dominant paraffin section IHC pattern. |
| Situation | Likely cause | Next action |
|---|---|---|
| A known High tissue gives little or no stain. | The result conflicts with HPA's reported High staining in cardiomyocytes and kidney tubular cells; the supplied sources do not identify a target-specific failure mechanism (HPA tissue IHC). | Check section integrity, positive control performance, retrieval settings and detection reagents, then repeat under documented IHC conditions (general IHC practice). Do not infer a target-specific fixation effect from this result. |
| Signal is diffuse across tissue and blank spaces. | The distribution differs from HPA's granular cytoplasmic pattern (HPA tissue IHC). Nonspecific binding or detection background is possible (general IHC practice). | Inspect a no-primary control, review blocking and washing, and check whether the detection chemistry deposits color without primary antibody (general IHC practice). Score cells only after background is controlled. |
| Unexpected cells stain strongly, including adipocytes. | HPA lists adipocytes as Not detected; apparent signal may reflect cell misidentification, cross-reactivity or endogenous chromogenic activity (HPA tissue IHC; general IHC practice). | Compare adjacent morphology and known positive cells; examine a no-primary control and, for enzyme-based detection, an appropriate endogenous activity control (general IHC practice). |
| The section is mainly nuclear rather than granular cytoplasmic. | HPA tissue IHC favors granular cytoplasm, although HPA ICC-IF also reports approved nucleoplasmic localization (HPA tissue IHC; HPA subcellular ICC-IF). | Review morphology, controls and antibody validation before assigning the nuclear signal to OPA1 (general IHC practice). Record the compartment separately from cytoplasmic IHC scoring. |
| Staining is weak in skeletal muscle myocytes or cerebral cortex endothelial cells. | HPA reports Low staining in these cell populations (HPA tissue IHC). A weak result there is less informative about assay performance than a reported High cell population. | Compare with a documented High population, such as heart cardiomyocytes or kidney tubular cells, and assess background before changing assay settings (HPA tissue IHC; general IHC practice). |
| Q: IF/ICC shows nucleoplasmic signal. Is that compatible with OPA1? | A: HPA lists nucleoplasm as approved and mitochondria as supported in ICC-IF, while tissue IHC is described as granular cytoplasmic (HPA subcellular ICC-IF; HPA tissue IHC). | Interpret the IF/ICC result using its own controls and guide; retain granular cytoplasmic staining as the primary paraffin section IHC expectation (HPA tissue IHC; general IHC practice). |
Experiment evidence
IHC & IF images showing OPA1 staining
PB9731 · IHCIHC analysis of OPA1 using anti-OPA1 antibody (PB9731). OPA1 was detected in paraffin-embedded section of Mouse Liver Tissue. Heat mediated
PB9731 · IHCIHC analysis of OPA1 using anti-OPA1 antibody (PB9731). OPA1 was detected in paraffin-embedded section of Rat Kidney Tissue. Heat mediated a
PB9731 · IFIF analysis of OPA1 using anti-OPA1 antibody (PB9731). OPA1 was detected in immunocytochemical section of U20S cells. Enzyme antigen retriev
A00508-1 · IFIF analysis of OPA1 using anti-OPA1 antibody (A00508-1). OPA1 was detected in an immunocytochemical section of U2OS cells. Enzyme antigen re
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000198836-OPA1/tissue/cerebral+cortex#imid_8978114
HPA · Salivary glandHuman Protein Atlas IHC, Salivary gland. Source: https://www.proteinatlas.org/ENSG00000198836-OPA1/tissue/salivary+gland#imid_8978062
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000198836-OPA1/tissue/colon#imid_8958080
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000198836-OPA1/tissue/liver#imid_8958081
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA036927; green = OPA1, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA036927; green = OPA1, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for OPA1 IHC & IF
🧪Run kidney first: OPA1 should stain tubular cells (HPA: High in kidney cells in tubules). Run adipose tissue as the negative comparator (HPA: Not detected in adipocytes); on the kidney slide, internal negative cells should show counterstain without specific DAB signal, but identify them empirically because the supplied HPA row does not establish a negative kidney cell type.
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 OPA1 in A-431, U-251MG, U2OS, with annotated localisation: Nucleoplasm (approved), Mitochondria (supported) (HPA subcellular).
Technical controls: Include no-primary (secondary-only) and concentration-matched rabbit IgG isotype controls, plus OPA1-knockout tissue if available; the selected IHC caption uses a rabbit primary antibody (PB9731 tissue-IHC caption). Block endogenous peroxidase and check for endogenous biotin in kidney when using the caption’s biotin-based DAB detection system (PB9731 tissue-IHC caption).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected caption does not state its fixative (PB9731 tissue-IHC caption). Its paraffin-section example uses heat retrieval in EDTA at pH 8.0, but whether retrieval is required for kidney is unreported (PB9731 tissue-IHC caption); neither frozen sections nor IF can be called easier from the supplied evidence. Kidney’s endogenous biotin may add background with the caption’s biotin-based detection system (PB9731 tissue-IHC caption).
HPA tissue IHC evidence for OPA1
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Supported — 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 |
|---|---|---|---|---|
| Appendix | Glandular cells | High | Protein (IHC) | HPA → |
| Duodenum | Glandular cells | High | Protein (IHC) | HPA → |
| Heart muscle | Cardiomyocytes | High | Protein (IHC) | HPA → |
| Kidney | Cells in tubules | High | Protein (IHC) | HPA → |
| Ovary | Follicle cells | High | 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 OPA1 IHC Tips
Use the catalog antibody’s paraffin-section result as the starting point, then assess OPA1 staining against its mitochondrial biology and tissue controls.
Which retrieval conditions should I start with for paraffin-section OPA1 IHC?
Start with heat-mediated retrieval in EDTA at pH 8.0 for paraffin sections (PB9731 tissue-IHC caption). The same caption reports OPA1 detection in mouse liver after this retrieval, followed by 10% goat-serum blocking and 1 μg/ml primary antibody overnight at 4°C (PB9731 tissue-IHC caption). If staining is weak, check heating consistency, section adhesion and antibody incubation before testing another retrieval buffer as a fallback (standard IHC practice). Keep retrieval time and temperature identical across comparison sections, because unequal heating can change apparent staining intensity (standard IHC practice). The caption does not report a retrieval duration or fixative (PB9731 tissue-IHC caption).
How should I troubleshoot fixation-related changes in OPA1 staining?
The catalog image identifies paraffin-embedded mouse liver but does not state its fixative, so OPA1-specific fixation sensitivity is unknown (PB9731 tissue-IHC caption). Record fixative, fixation duration, section thickness and processing history for each specimen before comparing chromogenic intensity (standard IHC practice). If a specimen stains weakly, compare it with a similarly processed positive section using the documented EDTA pH 8.0 retrieval and 1 μg/ml primary incubation (PB9731 tissue-IHC caption; standard IHC practice). Examine morphology and staining together, since damaged sections can make a negative result hard to interpret (standard IHC practice). Do not assign a target-specific fixation effect without matched evidence (PB9731 tissue-IHC caption).
Where should convincing OPA1 chromogenic staining appear?
Expect predominantly granular cytoplasmic staining in tissue sections, consistent with the reported IHC profile and mitochondrial localisation (HPA tissue IHC; UniProt O60313 localisation). OPA1 occupies the mitochondrial inner membrane and intermembrane space; its listed transmembrane segment spans residues 97–113 (UniProt O60313 topology). Review signal within intact cells against their cytoplasm and nuclei, using morphology rather than DAB colour alone to assign compartment (standard IHC practice). HPA also reports nucleoplasm in subcellular imaging, so investigate reproducible nuclear staining with orthogonal controls instead of automatically dismissing it (HPA subcellular). Diffuse extracellular deposits or staining confined to section edges warrant a processing and background check (standard IHC practice).
Can this IHC stain distinguish long and short OPA1 forms?
Do not score the catalog IHC signal as a specific OPA1 form without an epitope map and form-specific validation (PB9731 tissue-IHC caption; UniProt O60313 processing). The listed long chain spans residues 88–960, whereas the short chain spans 195–960; both can therefore retain an epitope within their shared sequence (UniProt O60313 processing). OPA1 also has 6 listed isoforms, and the caption supplies no epitope coordinates or isoform selectivity (UniProt O60313 isoforms; PB9731 tissue-IHC caption). Compare sections processed and stained together when assessing intensity (standard IHC practice). Changes in DAB intensity alone cannot establish altered proteolytic processing or an L-OPA1:S-OPA1 ratio (UniProt O60313 function; standard IHC interpretation).
How can IF help assess an ambiguous OPA1 IHC pattern?
Use IF as a separate validation experiment, since the supplied catalog caption documents chromogenic IHC in paraffin sections and gives no IF conditions (PB9731 tissue-IHC caption). In heart muscle, multiplex OPA1 with a cardiomyocyte marker such as cardiac troponin T and a mitochondrial marker, using distinct channels to assess cell identity and localisation (HPA tissue IHC: High in cardiomyocytes; standard IF practice). Choose a far-red OPA1 fluorophore when tissue autofluorescence compromises shorter-wavelength channels, and include single-label controls (standard IF practice). Permeabilisation must allow antibody access to the intermembrane-space epitope if that is the mapped epitope; optimise it without assuming the antibody’s epitope location (UniProt O60313 topology; standard IF practice).
What should I check when OPA1 DAB staining looks diffuse?
Compare a no-primary control with the stained section to identify signal from detection reagents or endogenous tissue activity (standard IHC practice). The documented workflow uses biotinylated secondary antibody, a streptavidin–biotin complex and DAB, so assess background from that detection sequence separately from primary-dependent staining (PB9731 tissue-IHC caption; standard IHC practice). Confirm that peroxidase blocking was effective and that blocking, washes and chromogen development were consistent across sections (standard IHC practice). The catalog image used 10% goat serum and 1 μg/ml primary antibody overnight at 4°C (PB9731 tissue-IHC caption). Retain granular cytoplasmic signal as the pattern to assess after background is controlled (HPA tissue IHC).
How should I quantify OPA1 staining across tissue sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and tissue region before scoring, then apply the same threshold and imaging settings to all slides (standard IHC practice). An H-score combines the percentage of cells at each intensity grade, while percentage-positive cells or positive area per mm² can answer narrower questions (standard IHC practice). Normalise counts to the number of eligible intact cells, or positive area to the measured viable tissue area, and report the denominator (standard IHC practice). Score granular cytoplasmic staining separately from nuclear or diffuse background because HPA describes the tissue pattern as cytoplasmic and granular (HPA tissue IHC). Compare like cell populations across specimens; HPA reports high cardiomyocyte staining but no detection in adipocytes (HPA tissue IHC).
How do I distinguish true OPA1 staining from artefact?
Treat granular cytoplasmic staining in intact cells as the primary pattern to evaluate, given OPA1’s mitochondrial location and the HPA tissue-IHC profile (UniProt O60313 localisation; HPA tissue IHC). Check cell identity against the expected specimen: HPA reports high staining in kidney tubular cells and cardiomyocytes, but no detection in adipocytes (HPA tissue IHC). Exclude section-edge deposits, necrotic areas and signal reproduced in a no-primary control from positive-cell scoring (standard IHC practice). Investigate strong uniform nuclear DAB signal or unexpected extracellular colour with matched controls; HPA subcellular imaging also lists nucleoplasm, so compartment alone is not definitive (HPA subcellular; standard IHC practice). Control endogenous peroxidase before interpreting DAB as antibody-dependent signal (standard IHC practice).
Boster reagents
Best OPA1 / Dynamin-like GTPase OPA1, mitochondrial IHC Antibodies
The catalog shows OPA1 IHC in paraffin sections from mouse liver, rat kidney, and human lung cancer (PB9731 image captions), plus IF in U2OS cells (A00508-1 image caption).
Real IHC data
Anti-OPA1 Antibody ®
Cat # PB9731
Real IF data
Anti-OPA1 Antibody ®
Cat # A00508-1
PB9731 will render with mouse liver paraffin-section IHC; its other IHC captions show rat kidney and human lung cancer (catalog image captions). A00508-1 will render with IF in U2OS cells and is listed for IF and ICC (catalog image caption; catalog applications).
Which to pick: Choose PB9731 for tissue IHC and cross-species work: its own paraffin-section captions cover mouse, rat, and human samples, using 1 μg/ml primary antibody after EDTA pH 8.0 retrieval (PB9731 image captions). Choose A00508-1 for IF/ICC: its own U2OS image documents IF at 5 μg/ml, and ICC and IF are listed applications (A00508-1 image caption; catalog applications). Neither card reports a clone, and the PB9731 paraffin-section captions do not report a fixative (catalog clone fields; PB9731 image captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry O60313 (OPA1_HUMAN, Dynamin-like GTPase OPA1, mitochondrial).
- Human Protein Atlas. OPA1 tissue IHC expression (reliability: Supported).
- Human Protein Atlas. OPA1 subcellular location (ICC-IF): Mainly localized to the nucleoplasm and mitochondria..
- Human Protein Atlas. OPA1 antibody validation summary (2 antibodies).
- Fusion or Fission: The Destiny of Mitochondria In Traumatic Brain Injury of Different Severities. Scientific reports 2017 — PMC5569027.
- SS31 Alleviates Pressure Overload-Induced Heart Failure Caused by Sirt3-Mediated Mitochondrial Fusion. Frontiers in cardiovascular medicine 2022 — PMC9110818.
- Lycium Barbarum Polysaccharides Decrease Hyperglycemia-Aggravated Ischemic Brain Injury through Maintaining Mitochondrial Fission and Fusion Balance. International journal of biological sciences 2017 — PMC5555107.
- Mitochondrial role in adaptive response to stress conditions in preeclampsia. Scientific reports 2016 — PMC5004102.
- PubMed PMID:9628581 — UniProt-cited evidence.
- PubMed PMID:20843780 — UniProt-cited evidence.
- PubMed PMID:16641997 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for OPA1 (Dynamin-like GTPase OPA1, mitochondrial). Expected localisation Granular cytoplasm (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet PB9731). Validated IHC antibody: PB9731 at 0.5-1μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Appendix (Glandular cells); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded Mouse Liver tissue; fixative not specified (datasheet PB9731); 2. Fixation: Image fixative and duration unreported (datasheet PB9731); 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 PB9731); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet PB9731); 8. Primary antibody: Rabbit anti-OPA1, 0.5-1μg/ml (datasheet PB9731); 9. Primary incubation: Overnight at 4 °C (datasheet PB9731). Watch for: Fixation: Keep fixation consistent across sections. (standard IHC practice; not target-specific); Caveat: Staining has medium concordance with RNA (HPA tissue IHC); Isoform / epitope: Six isoforms; long and cleaved short forms may differ at an unknown epitope (UniProt); Regulation: Low tissue specificity (HPA tissue RNA). Evidence: UniProt O60313, HPA tissue IHC · Supported, HPA ICC-IF · Mainly localized to the nucleoplasm and mitochondria., Datasheet PB9731, PMC9110818, PMC5555107. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/OPA1/immunohistochemistry-ihc-guide.