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ACAA2 / 3-ketoacyl-CoA thiolase, mitochondrial · IHC design guide
Design Immunohistochemistry for ACAA2
Plan chromogenic ACAA2 IHC in paraffin sections using catalog antibody M08341-1 at 1:50 (datasheet M08341-1). Expect granular cytoplasmic staining in hepatocytes and kidney tubule cells, and include controls because staining and RNA show low consistency (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
ACAA2 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); mitochondrion (UniProt) | |
| Staining pattern | Granular cytoplasm in hepatocytes and kidney tubule cells (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet M08341-1) | |
| 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 | Keep fixation consistent across paraffin sections. (standard IHC practice; not target-specific) | |
| Caveat | Staining and RNA show low consistency; verify independently (HPA tissue IHC) | |
| Regulation | Liver-enhanced RNA expression (HPA tissue RNA) | |
| Isoform / epitope | No annotated isoforms; chain spans residues 1–397 (UniProt) |
Section 1
Recommended ACAA2 IHC & IF Protocols
The catalog antibody protocol (datasheet: M08341-1) is accompanied by two published ACAA2 IHC protocols (PMC12771824; PMC11940541).
| Sample | Paraffin-embedded human colorectal adenocarcinoma tissue; fixative not specified (datasheet M08341-1) |
| Fixation | Image fixative and duration unreported (datasheet M08341-1); 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 M08341-1); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet M08341-1) |
| Primary antibody | Rabbit monoclonal (clone 26A90) anti-ACAA2, 1:50 (datasheet M08341-1) |
| Primary incubation | Overnight at 4 °C (datasheet M08341-1) |
| Detection | HRP-conjugated secondary, DAB chromogen (datasheet M08341-1) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | ACAA2-positive staining in glandular cells of appendix (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression with granular pattern in several tissues. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA retrieval at pH 8.0 (datasheet: M08341-1); citrate retrieval at pH 6.0 is a published alternative (PMC12771824: IHC methods).
Section 2
What Is the Expected ACAA2 Staining Pattern?
ACAA2 is mitochondrial and has no transmembrane segment (UniProt P42765). In paraffin-section IHC, expect granular cytoplasmic staining in hepatocytes, kidney tubule cells, and several glandular epithelia (HPA tissue IHC). HPA rates its tissue IHC evidence Approved, but reports low consistency with RNA data and pending external verification (HPA tissue IHC).
| Granular cytoplasmic signal in hepatocytes or kidney tubule cells. | This matches cells rated High by HPA and its granular cytoplasmic tissue profile (HPA tissue IHC). The pattern is consistent with mitochondrial ACAA2 (UniProt P42765); chromogenic granules alone do not prove organelle identity. |
| Predominantly nuclear staining, with little granular cytoplasmic signal. | A nuclear dominant pattern conflicts with the mitochondrial assignment (UniProt P42765) and HPA tissue profile (HPA tissue IHC). Check staining controls and antibody specificity before scoring it as ACAA2. |
| Strong staining in adipocytes or esophageal squamous epithelial cells. | HPA reports ACAA2 as Not detected in those cell types (HPA tissue IHC). Cross-reactivity or endogenous detection activity is possible; investigate with detection controls and a known-positive section (standard IHC practice). |
| Diffuse color across tissue and empty slide areas. | This does not resemble HPA's granular cytoplasmic profile (HPA tissue IHC). Background from the staining workflow is possible; review blocking, washes, and the detection-only control (standard IHC practice). |
| No signal in hepatocytes on an otherwise assessable liver section. | HPA rates hepatocyte staining High, so this result needs investigation (HPA tissue IHC). Check section quality, retrieval and detection controls, and antibody dilution before interpreting absence as biological (standard IHC practice). |
💡Expected ACAA2 appearanceCall a section positive when hepatocytes, kidney tubule cells, or the listed glandular cells show clear granular cytoplasmic staining, expected to be High by HPA; dominant nuclear staining or strong signal in HPA Not detected cell types is suspect (HPA tissue IHC; UniProt P42765).
| Compartment and topology | Mitochondrial location and absence of a transmembrane segment support an intracellular pattern (UniProt P42765). HPA describes tissue staining as granular cytoplasmic (HPA tissue IHC). |
| Choice of comparison tissue | Liver hepatocytes and kidney tubule cells are High, whereas adipocytes and esophageal squamous epithelial cells are Not detected (HPA tissue IHC). Compare the specified cell populations, not whole-section averages. |
| Strength of tissue evidence | The tissue IHC rating is Approved, with low agreement between antibody staining and RNA expression and external verification pending (HPA tissue IHC). Treat unexpected intensity as a finding to verify, not a definitive expression change. |
| IF/ICC cross-check | Where should IF/ICC signal appear? HPA supports mitochondria as the main location; cytosol and basal body are uncertain additional locations (HPA subcellular ICC-IF). This answers localisation only and supplies no IF protocol. |
| Antigen retrieval | Target-specific fixation effects are not established by the supplied assay evidence. Verify with a matched IHC source before attributing a result to fixation. |
| Situation | Likely cause | Next action |
|---|---|---|
| Liver positive control has no hepatocyte staining. | Hepatocytes are rated High (HPA tissue IHC); an absent signal may reflect a staining-workflow failure (standard IHC practice). | Confirm section quality and control performance; review retrieval, antibody dilution, and detection steps before scoring the sample (standard IHC practice). |
| Signal is mainly nuclear. | This conflicts with mitochondrial ACAA2 (UniProt P42765) and granular cytoplasmic tissue staining (HPA tissue IHC). | Compare a known-positive section and detection-only control; reassess antibody specificity if the nuclear pattern persists (standard IHC practice). |
| Adipocytes stain strongly. | Adipocytes are Not detected in HPA tissue IHC; cross-reactivity or endogenous detection activity may account for signal (HPA tissue IHC; standard IHC practice). | Check the detection-only control and compare with a High-rated cell population on a separately processed section (HPA tissue IHC; standard IHC practice). |
| Color is diffuse across cells and background areas. | The appearance differs from HPA's granular cytoplasmic profile (HPA tissue IHC); nonspecific background is possible (standard IHC practice). | Review blocking, washes, and detection-only controls; score cellular granules only when distinguishable from background (standard IHC practice). |
| Kidney staining is present, but the cell population is unclear. | HPA's High call applies to cells in tubules, not every kidney cell (HPA tissue IHC). | Identify tubules using tissue morphology and the counterstain, then score the relevant cells separately (HPA tissue IHC; standard IHC practice). |
| Tissue intensity differs from the HPA example. | HPA reports low consistency between antibody staining and RNA expression, with external verification pending (HPA tissue IHC). Technical variation is also possible (standard IHC practice). | Compare matched control sections and cell-specific patterns; document the difference rather than assigning a biological cause from intensity alone (standard IHC practice). |
Experiment evidence
IHC & IF images showing ACAA2 staining
M08341-1 · IHCIHC analysis of ACAA2 using anti-ACAA2 antibody (M08341-1). ACAA2 was detected in a paraffin-embedded section of human colorectal adenocarci
M08341-1 · IHCIHC analysis of ACAA2 using anti-ACAA2 antibody (M08341-1). ACAA2 was detected in a paraffin-embedded section of human colung cancer tissue.
M08341-1 · IFIF analysis of ACAA2 using anti-ACAA2 antibody (M08341-1). ACAA2 was detected in an immunocytochemical section of Hela cells. Enzyme antigen
PB10022 · IHCIHC analysis of ACAA2 using anti-ACAA2 antibody (PB10022). ACAA2 was detected in paraffin-embedded section of human intestinal cancer tissue
PB10022 · IFIF analysis of ACAA2 using anti-ACAA2 antibody (PB10022). ACAA2 was detected in immunocytochemical section of U20S cell. Enzyme antigen retr
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000167315-ACAA2/tissue/cerebral+cortex#imid_11124423
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000167315-ACAA2/tissue/colon#imid_11122397
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000167315-ACAA2/tissue/liver#imid_11122398
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000167315-ACAA2/tissue/kidney#imid_11122415
HPA · ICC-IF CACO-2Human Protein Atlas immunofluorescence, CACO-2, antibody HPA042303; green = ACAA2, blue = nucleus, red = microtubules.
HPA · ICC-IF CACO-2Human Protein Atlas immunofluorescence, CACO-2, antibody HPA042303; green = ACAA2, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for ACAA2 IHC & IF
🧪Run colon first and expect glandular cells to stain (HPA: High in colon glandular cells). Use cervix glandular cells as the negative tissue (HPA: Not detected in cervix glandular cells); on the colon slide, assess adjacent nonglandular cells for background without assuming they are ACAA2-negative.
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 ACAA2 in CACO-2, Hep-G2, U2OS, ASC52telo, RPTEC/TERT1, hTERT-RPE1 (serum starved), NIH 3T3, with annotated localisation: Mitochondria (supported) (HPA subcellular).
Technical controls: Include no-primary (secondary-only), host- and isotype-matched rabbit IgG controls, and ACAA2-knockout tissue as a biological negative (caption: rabbit anti-ACAA2 primary; standard IHC practice). Block endogenous peroxidase and check for DAB background in colon sections (standard chromogenic IHC practice; HPA: High in colon glandular cells).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected M08341-1 paraffin-section caption does not state the fixative (selected-SKU caption). The caption reports heat retrieval in EDTA at pH 8.0 and primary antibody at 1:50 overnight at 4°C, but does not establish that retrieval is required (selected-SKU caption). HPA supports mitochondrial localization in ICC-IF images, but the supplied evidence does not establish whether frozen sections or IF are easier than paraffin IHC; assess colon sections for endogenous peroxidase background during chromogenic detection (HPA: mitochondria supported; standard IHC practice).
HPA tissue IHC evidence for ACAA2
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — 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 |
|---|---|---|---|---|
| Appendix | Glandular 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 → |
| Kidney | Cells in tubules | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Cerebellum | Cells in granular layer | Not detected | Protein (IHC) | HPA → |
| Cervix | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Esophagus | Squamous epithelial cells | Not detected | Protein (IHC) | HPA → |
| Lymph node | Germinal center cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced ACAA2 IHC Tips
Troubleshoot ACAA2 staining in paraffin sections by checking retrieval, signal location, controls, and scoring against the documented tissue pattern.
How should I retrieve ACAA2 when chromogenic staining is weak?
Start with heat-mediated retrieval in EDTA pH 8.0 for paraffin sections (datasheet M08341-1). The documented colorectal adenocarcinoma staining used this retrieval, followed by 10% goat serum block and 1:50 primary antibody overnight at 4°C (datasheet M08341-1). If signal remains weak, compare controlled changes in heating duration while keeping section thickness, antibody incubation, and DAB development constant (standard IHC practice). Assess whether granular cytoplasmic signal improves in expected cells without a parallel rise in diffuse background, using a known positive section in each run (HPA: granular cytoplasmic tissue pattern; standard IHC practice).
Could fixation explain inconsistent ACAA2 staining between paraffin blocks?
The selected paraffin-section caption does not state a fixative, so ACAA2 sensitivity to fixation cannot be determined from that experiment (datasheet M08341-1). Record each block’s fixative and fixation interval, then compare sections processed together with the same EDTA pH 8.0 retrieval and antibody incubation (datasheet M08341-1; standard IHC practice). Underfixation and prolonged fixation can each alter morphology or epitope accessibility in routine IHC, so examine tissue preservation alongside staining intensity (standard IHC practice). Do not infer an ACAA2-specific fixation effect from its mitochondrial location, modified residues, or tissue staining pattern (UniProt P42765; HPA: tissue IHC profile).
What cellular staining pattern should count as plausible ACAA2 signal?
Look for granular cytoplasmic staining, consistent with ACAA2’s mitochondrial location and the reported tissue IHC pattern (UniProt P42765: mitochondrion; HPA: granular cytoplasmic expression). Supported subcellular imaging places ACAA2 mainly in mitochondria, whereas cytosol and basal body assignments remain uncertain (HPA: subcellular location). A uniformly nuclear or surface-membrane signal should prompt review of antibody specificity, counterstain overlap, and nonspecific chromogen deposition before scoring (UniProt P42765: mitochondrial location and no transmembrane segment; standard IHC practice). Compare the pattern within intact cells at matched magnification and development time, including a no-primary control to identify detection-system deposits (standard IHC practice).
How should I investigate an unexpected ACAA2 epitope-dependent IHC pattern?
The record lists 0 isoforms and a single annotated chain spanning residues 1–397, so an alternative annotated isoform does not explain discordant staining (UniProt P42765). ACAA2 has no annotated transmembrane segment, while multiple modified residues include acetylation, succinylation, and phosphorylation sites (UniProt P42765). Without the catalog antibody’s mapped epitope, do not attribute a staining difference to any particular modification or residue (UniProt P42765: modified residues; datasheet M08341-1: epitope not supplied). Compare independently validated antibodies against distinct documented epitopes, where available, on adjacent sections with identical retrieval and detection conditions (standard IHC practice).
How can IF help resolve ambiguous ACAA2 staining in tissue?
Use IF as a separate corroboration experiment, pairing ACAA2 with a marker for the expected cell type and a mitochondrial marker to assess cellular identity and subcellular overlap (HPA: tissue cell patterns and supported mitochondrial location; standard IF practice). Choose spectrally separated fluorophores after checking tissue autofluorescence, and include single-label controls when interpreting multiplexed channels (standard IF practice). Because ACAA2 has no transmembrane segment and is mainly mitochondrial, optimise permeabilisation to allow antibody access to its intracellular epitope; the epitope’s exact position is not supplied (UniProt P42765; HPA: subcellular location; standard IF practice). Keep IF fixation and permeabilisation conclusions separate from the paraffin IHC caption, which does not report a fixative (datasheet M08341-1).
How do I reduce diffuse brown staining without losing ACAA2 signal?
Compare stained sections with no-primary and secondary-only controls to distinguish antibody-associated background from deposits produced by the detection workflow (standard IHC practice). Block endogenous peroxidase before HRP and DAB detection, and verify that the block itself preserves tissue morphology (standard IHC practice; datasheet M08341-1: HRP and DAB detection). The documented section used 10% goat serum and 1:50 rabbit primary overnight at 4°C; use those settings as a traceable starting point for this SKU (datasheet M08341-1). If background persists, titrate primary concentration and DAB development together with a positive control, judging improvement by granular cellular contrast (standard IHC practice; HPA: granular cytoplasmic pattern).
How should I score ACAA2 across sections with different cell composition? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring; count granular cytoplasmic ACAA2 signal within comparable viable cells (HPA: granular cytoplasmic pattern; UniProt P42765: mitochondrial location; standard IHC practice). Report the percentage of positive cells and an H-score based on the proportion at each intensity, using the same thresholds and imaging settings across sections (standard IHC practice). Normalise each result to the number of evaluable cells in that cell type, or report positive-cell density per mm² of viable tissue when area is the denominator (standard IHC practice). Keep epithelial and stromal compartments separate where relevant, and exclude folded, necrotic, or edge-damaged regions consistently (standard IHC practice).
When is an apparent ACAA2-positive region likely to be artefactual?
A credible result has granular cytoplasmic signal in intact cells and fits ACAA2’s supported mitochondrial localisation (HPA: tissue IHC pattern and supported subcellular location; UniProt P42765). High staining in liver hepatocytes or kidney tubule cells is a useful comparison, while a strong signal in a population reported as undetected warrants extra verification (HPA: tissue IHC levels). Treat staining confined to section edges, necrotic areas, or no-primary controls as suspect, and check endogenous peroxidase before interpreting DAB deposits (standard IHC practice). HPA tissue staining has approved reliability but low consistency with RNA data and awaits external verification, so confirm consequential findings independently (HPA: reliability description).
Boster reagents
Best ACAA2 / 3-ketoacyl-CoA thiolase, mitochondrial IHC Antibodies
Both anti-ACAA2 antibodies have IHC images from human paraffin sections and IF images from cultured cells (catalog image captions); both list Human, Mouse and Rat reactivity (catalog reactivity).
Real IHC data
Anti-ACAA2 Rabbit Monoclonal Antibody
Cat # M08341-1
Real IHC data
Anti-ACAA2 Antibody ®
Cat # PB10022
M08341-1 shows IHC staining in a human colorectal adenocarcinoma paraffin section and IF staining in HeLa cells (M08341-1 image captions). PB10022 shows IHC staining in a human intestinal cancer paraffin section and IF staining in U20S cells (PB10022 image captions).
Which to pick: For paraffin-section IHC, choose M08341-1 for a monoclonal antibody with a documented 1:50 IHC example, or PB10022 for an antibody with a documented 1 μg/ml IHC example (catalog clone; each SKU’s IHC caption). For IF/ICC, both list those applications and have cell-based IF images; M08341-1 is monoclonal, while PB10022 has no clone specified (catalog applications and clone; IF captions). Both list Human, Mouse and Rat reactivity, but their shown IHC examples are human paraffin sections, and neither IHC caption reports the fixative (catalog reactivity; IHC captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P42765 (THIM_HUMAN, 3-ketoacyl-CoA thiolase, mitochondrial).
- Human Protein Atlas. ACAA2 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. ACAA2 subcellular location (ICC-IF): Mainly localized to the mitochondria. In addition localized to the cytosol and basal body..
- Human Protein Atlas. ACAA2 antibody validation summary (1 antibodies).
- The metabolic crossroads: glycolysis in cancer and neurodegeneration. BMC neurology 2025 — PMC12771824.
- Acetyl-CoA acyltransferase 2 palmitoylation drives liver fibrosis by inducing hepatic stellate cell ferroptosis. Redox biology 2026 — PMC12859212.
- Metabolic classifications of renal cell carcinoma reveal intrinsic connections with clinical and immune characteristics. Journal of translational medicine 2023 — PMC9960222.
- ACAA2 Protects Against Cardiac Dysfunction and Lipid Peroxidation in Renal Insufficiency with the Treatment of S-Nitroso-L-Cysteine. Biomolecules 2025 — PMC11940541.
- PubMed PMID:8241273 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
- PubMed PMID:15242332 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for ACAA2 (3-ketoacyl-CoA thiolase, mitochondrial). Expected localisation Granular cytoplasm (HPA tissue IHC); mitochondrion (UniProt). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet M08341-1). Validated IHC antibody: M08341-1 at 1:50 (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 human colorectal adenocarcinoma tissue; fixative not specified (datasheet M08341-1); 2. Fixation: Image fixative and duration unreported (datasheet M08341-1); 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 M08341-1); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet M08341-1); 8. Primary antibody: Rabbit monoclonal (clone 26A90) anti-ACAA2, 1:50 (datasheet M08341-1); 9. Primary incubation: Overnight at 4 °C (datasheet M08341-1). Watch for: Fixation: Keep fixation consistent across paraffin sections. (standard IHC practice; not target-specific); Caveat: Staining and RNA show low consistency; verify independently (HPA tissue IHC); Isoform / epitope: No annotated isoforms; chain spans residues 1–397 (UniProt); Regulation: Liver-enhanced RNA expression (HPA tissue RNA). Evidence: UniProt P42765, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the mitochondria. In addition localized to the…, Datasheet M08341-1, PMC12771824, PMC11940541. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/ACAA2/immunohistochemistry-ihc-guide.