ACADVL / Very long-chain acyl-CoA dehydrogenase, mitochondrial · IHC design guide

Design Immunohistochemistry for ACADVL

This guide covers paraffin-section ACADVL IHC with the catalog antibody at 2–5 μg/ml (datasheet PB10023). It uses strongly stained cardiomyocytes and unstained chondrocytes as tissue references for interpreting granular cytoplasmic staining (HPA tissue IHC).

Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use

Immunohistochemistry protocol sheet for ACADVL (IHC for ACADVL): expected localisation Granular cytoplasm (HPA tissue IHC); mitochondrial inner membrane (UniProt), antibody PB10023, validated IHC image, and IHC protocol steps — Printable **ACADVL** IHC protocol sheet — expected localisation Granular cytoplasm (HPA tissue IHC); mitochondrial inner membrane (UniProt), antibody PB10023, controls and protocol steps. Open the full ACADVL IHC guide →

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ACADVL 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); mitochondrial inner membrane (UniProt)
	Staining pattern	Granular cytoplasmic staining in cardiomyocytes (HPA tissue IHC)
	Antigen retrieval	EDTA pH 8.0 HIER, heat-mediated (datasheet PB10023)
	Positive control ⓘ	Adrenal gland+4 more · see all
	Negative control ⓘ	Soft tissue

Important caveats

Reasons your staining may differ from the expected pattern.

	Fixation	Keep fixation consistent across sections (standard IHC practice; not target-specific); Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image PB10023)
	Caveat	Strongly stained lung macrophages can affect cell-type scoring (HPA tissue IHC)
	Regulation	Enriched in heart and skeletal muscle (UniProt)
	Isoform / epitope	3 isoforms; mature chain 41–655; epitope impact unknown (UniProt)

Section 1

Recommended ACADVL IHC & IF Protocols

The catalog antibody uses heat retrieval with EDTA at pH 8.0 (datasheet: PB10023). One published ACADVL IHC protocol describes staining of GM tissue sections (PMC11308078).

Recommended immunohistochemistry (IHC-P) protocol parameters
Sample	Paraffin-embedded human liver cancer tissue; fixative not specified (datasheet PB10023)
Fixation	Image fixative and duration unreported (datasheet PB10023); 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 PB10023); 20 min, 95–100 °C (standard)
Peroxidase block	3% H2O2, 10 min, room temperature (standard)
Blocking	10% goat serum (datasheet PB10023)
Primary antibody	Rabbit anti-ACADVL, 2-5 μg/ml (datasheet PB10023)
Primary incubation	Overnight at 4 °C (datasheet PB10023)
Detection	HRP-conjugated secondary, DAB chromogen (datasheet PB10023)
Counterstain	Hematoxylin, blue, dehydrate and mount (standard)
Expected result	ACADVL-positive staining in glandular cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: General cytoplasmic expression with a granular pattern. No signal in the no-primary control.

💡Decision noteStart with heat retrieval using EDTA at pH 8.0 for the catalog antibody (datasheet: PB10023). The published protocol does not specify its retrieval buffer or pH (PMC11308078).

Section 2

What Is the Expected ACADVL Staining Pattern?

ACADVL is assigned to the mitochondrial inner membrane without a transmembrane segment (UniProt P49748 topology). In paraffin sections, expect granular cytoplasmic staining (HPA: tissue IHC profile), especially in cardiomyocytes and several glandular cell populations (HPA: High). HPA rates the tissue IHC evidence Enhanced, with medium consistency between antibody staining and RNA expression (HPA: reliability). Interpret intensity alongside cell identity and staining controls.

What am I looking at on my slide?
Granular cytoplasmic staining in cardiomyocytes; similar staining in adrenal or colon glandular cells (HPA: High in each cell type).	This matches HPA tissue IHC and the mitochondrial location assigned by UniProt (HPA: granular cytoplasmic profile; UniProt P49748 location). Compare the signal with local background before calling a cell positive (general IHC practice).
Strong, uniform nuclear staining with little granular cytoplasmic signal.	This differs from the expected tissue IHC pattern (HPA: granular cytoplasmic profile). Review controls and antibody dilution before interpreting it as ACADVL (general IHC practice). HPA reports uncertain nucleoplasmic and nucleolar signals in ICC-IF, so nuclear signal alone is inconclusive (HPA: additional locations uncertain).
Prominent staining of chondrocytes in soft tissue, especially when other cells show little signal.	HPA reports ACADVL as not detected in these chondrocytes (HPA: soft-tissue chondrocytes Not detected). Consider nonspecific antibody binding or endogenous detection activity; neither can be distinguished from the image alone (general IHC practice).
Diffuse chromogen over tissue and blank areas, obscuring cell boundaries and granules.	This is difficult to score against the expected granular cytoplasmic pattern (HPA: tissue IHC profile). Check primary-omission and detection controls, then review blocking, washes and chromogen development (general IHC practice).
No convincing staining in a heart section with identifiable cardiomyocytes.	Cardiomyocytes are an HPA High cell population, making this a useful run-level warning (HPA: heart-muscle cardiomyocytes High). Check the section and detection controls, then verify the antibody's IHC-P instructions; absence of signal alone does not establish absent ACADVL (general IHC practice).

💡Expected ACADVL appearanceCall a positive result when identifiable cardiomyocytes or HPA High glandular cells show distinct granular cytoplasmic signal above background (HPA: tissue IHC profile and High cell populations); diffuse chromogen or dominant nuclear-only staining should prompt control review before scoring (general IHC practice; HPA: nucleoplasmic location uncertain).

How each factor affects the staining
Cell population chosen for comparison	Heart-muscle cardiomyocytes, adrenal and colon glandular cells, and lung macrophages are reported High; soft-tissue chondrocytes are Not detected (HPA: tissue IHC). These are reference patterns, not an intensity guarantee for every section (general IHC practice).
Subcellular interpretation	UniProt assigns ACADVL to the mitochondrial inner membrane and lists no transmembrane segment (UniProt P49748 topology). HPA describes tissue staining as generally granular and cytoplasmic; light-microscope IHC cannot resolve the inner membrane itself (HPA: tissue IHC profile; general IHC practice).
Antibody evidence	HPA019006 and HPA020595 each have Enhanced IHC validation (HPA: antibody IHC status). HPA's tissue profile has medium consistency with RNA data, so use the reported cell pattern with controls rather than treating staining intensity as a quantitative protein measurement (HPA: reliability; general IHC practice).
Processing and isoforms	UniProt lists a mature chain spanning residues 41–655 and 3 isoforms (UniProt P49748 processing and isoforms). Without a supplied epitope map, their effect on this antibody's staining cannot be predicted; compare only results obtained with a documented IHC reagent (general IHC practice).
IF/ICC Q&A: Where should ACADVL fluorescence appear?	Mainly in mitochondria (HPA: ICC-IF supported main location). HPA also reports uncertain nucleoplasmic and nucleolar locations (HPA: ICC-IF additional locations). This IF/ICC observation helps interpret localization; the separate IF/ICC guide covers its workflow.

Why is my staining missing, weak or wrong?
Situation	Likely cause	Next action
Heart cardiomyocytes are blank while the rest of the run stains.	A failed local signal or section issue is possible; HPA reports High staining in cardiomyocytes (HPA: heart-muscle tissue IHC).	Check morphology, tissue coverage and run controls; verify the catalog antibody's IHC-P conditions before repeating the section (general IHC practice).
All sections, including expected positive cells, are blank.	A run-level detection or reagent problem is possible; heart cardiomyocytes are a reported High reference (HPA: tissue IHC).	Check the detection control, reagent order and expiration, then follow the documented IHC-P workflow for the antibody (general IHC practice).
The slide shows broad brown haze that hides granular signal.	Background from detection chemistry, inadequate washing or overly strong development is possible (general IHC practice).	Compare a primary-omission control, inspect blank tissue areas, and adjust blocking, washes or development within the detection system's instructions (general IHC practice).
Nuclei stain more strongly than cytoplasm.	The pattern conflicts with granular cytoplasmic tissue IHC; HPA's nuclear ICC-IF locations are uncertain (HPA: tissue IHC profile; ICC-IF additional locations).	Review counterstain and a primary-omission control; do not score nuclear-only chromogen as confirmed ACADVL (general IHC practice).
Chondrocytes stain strongly in soft tissue.	This conflicts with HPA's Not detected result for chondrocytes; nonspecific binding or endogenous detection activity is possible (HPA: soft-tissue IHC; general IHC practice).	Compare primary-omission and detection controls, then assess whether staining remains cell-associated and granular (general IHC practice; HPA: tissue IHC profile).
Signal is weak in one tissue but clear in another.	HPA reports different cell-level intensities, including Low in adipocytes and High in cardiomyocytes (HPA: tissue IHC).	Identify the stained cell population before changing conditions; compare like cells and confirm run quality with a reported High reference (general IHC practice; HPA: tissue IHC).

Source: UniProt P49748 Human Protein Atlas tissue Human Protein Atlas subcellular

Experiment evidence

IHC & IF images showing ACADVL staining

IHC analysis of ACADVL using anti-ACADVL antibody (PB10023). ACADVL was detected in a paraffin-embedded section of human liver cancer tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 2 μg/ml rabbit anti-ACADVL Antibody (PB10023) overnight at 4°C. Peroxidase Conjugated Goat Anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using HRP Conjugated Rabbit IgG Super Vision Assay Kit (Catalog # SV0002) with DAB as the chromogen.

PB10023 · IHCIHC analysis of ACADVL using anti-ACADVL antibody (PB10023). ACADVL was detected in a paraffin-embedded section of human liver cancer tissue

IHC analysis of ACADVL using anti-ACADVL antibody (PB10023). ACADVL was detected in a paraffin-embedded section of human ovarian cancer tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 2 μg/ml rabbit anti-ACADVL Antibody (PB10023) overnight at 4°C. Peroxidase Conjugated Goat Anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using HRP Conjugated Rabbit IgG Super Vision Assay Kit (Catalog # SV0002) with DAB as the chromogen.

PB10023 · IHCIHC analysis of ACADVL using anti-ACADVL antibody (PB10023). ACADVL was detected in a paraffin-embedded section of human ovarian cancer tiss

IF analysis of ACADVL using anti-ACADVL antibody (PB10023). ACADVL was detected in an immunocytochemical section of U2OS cells. Enzyme antigen retrieval was performed using IHC enzyme antigen retrieval reagent (AR0022) for 15 mins. The cells were blocked with 10% goat serum. And then incubated with 5 μg/mL rabbit anti-ACADVL Antibody (PB10023) overnight at 4°C. DyLight®488 Conjugated Goat Anti-Rabbit IgG (BA1127) was used as secondary antibody at 1:500 dilution and incubated for 30 minutes at 37°C. The section was counterstained with DAPI. Visualize using a fluorescence microscope and filter sets appropriate for the label used.

PB10023 · IFIF analysis of ACADVL using anti-ACADVL antibody (PB10023). ACADVL was detected in an immunocytochemical section of U2OS cells. Enzyme antig

HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000072778-ACADVL/tissue/cerebral+cortex#imid_5028684

HPA · Adrenal glandHuman Protein Atlas IHC, Adrenal gland. Source: https://www.proteinatlas.org/ENSG00000072778-ACADVL/tissue/adrenal+gland#imid_5028660

HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000072778-ACADVL/tissue/colon#imid_5028618

HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000072778-ACADVL/tissue/liver#imid_5028627

HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA019006; green = ACADVL, blue = nucleus, red = microtubules.

← swipe / scroll for more images →

Real IHC / IF data behind the staining-pattern expectations in this section.

Sample controls for ACADVL IHC & IF

🧪Run colon first: glandular cells should stain (HPA: High in colon glandular cells). Use soft tissue chondrocytes as the biological negative (HPA: Not detected); any unstained non-glandular cells on the colon slide should show counterstain without convincing granular cytoplasmic DAB signal, but are background comparators rather than validated ACADVL-negative cells (UniProt P49748: mitochondrial inner membrane; HPA: colon glandular cells High).

Positive control tissue: Adrenal gland (Glandular cells, HPA High)

Negative control tissue: Soft tissue (HPA Not detected)

ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show ACADVL in A-431, A-549, U2OS, Sperm, KOLF2.1J, NIH 3T3, with annotated localisation: Mitochondria (supported) (HPA subcellular).

Technical controls: Include a no-primary, secondary-only section, an isotype-matched rabbit IgG control, and an ACADVL knockout specimen where available (selected PB10023 caption: rabbit primary and anti-rabbit secondary; standard IHC practice). Quench endogenous peroxidase before HRP/DAB detection and compare any brown signal with the no-primary section (selected PB10023 caption: HRP/DAB; standard IHC practice).

⚠️Feasibility: No target-specific fixation window or fixation effect is reported in the supplied evidence; the selected PB10023 paraffin-section caption does not state its fixative. That caption reports heat retrieval in EDTA pH 8.0, but does not establish whether retrieval is required for every specimen (selected PB10023 caption). These data do not establish that frozen sections or IF are easier; in colon, check luminal debris and chromogen deposits against the no-primary control when scoring glandular-cell staining (HPA: colon glandular cells High; standard IHC practice).

HPA tissue IHC evidence for ACADVL

Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Enhanced — 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
Adrenal gland	Glandular cells	High	Protein (IHC)	HPA →
Bronchus	Ciliated cells (cell body)	High	Protein (IHC)	HPA →
Colon	Glandular cells	High	Protein (IHC)	HPA →
Duodenum	Glandular cells	High	Protein (IHC)	HPA →
Gallbladder	Glandular cells	High	Protein (IHC)	HPA →

Undetected expression · recommended negative controls

Tissue	Cell type	Level	Evidence	Source
Soft tissue	Chondrocytes	Not detected	Protein (IHC)	HPA →

Section 3

Advanced ACADVL IHC Tips

Troubleshoot ACADVL staining in paraffin sections by checking retrieval, granular cytoplasmic localisation, and cell-specific controls before comparing signal intensity.

What should I adjust when ACADVL staining is weak after retrieval?

Start with heat-mediated antigen retrieval in EDTA at pH 8.0 for paraffin sections (datasheet PB10023). The selected tissue image used this retrieval before overnight incubation with 2 µg/ml antibody at 4°C, so keep antibody concentration fixed while assessing retrieval (caption PB10023). Include heart muscle as a positive comparator and look for granular cytoplasmic staining in cardiomyocytes (HPA: High in cardiomyocytes; HPA: granular cytoplasmic profile). If signal remains weak, compare shorter and longer heating intervals on adjacent sections while retaining EDTA at pH 8.0 (standard IHC optimization practice).

How should I troubleshoot suspected fixation-related loss of ACADVL staining?

ACADVL-specific fixation sensitivity is unknown because the selected paraffin-section caption does not state a fixative (caption PB10023). Record the actual fixative and fixation duration for each specimen, then compare sections processed under those documented conditions using the same EDTA pH 8.0 retrieval (caption PB10023; standard IHC practice). If archival sections vary in staining, assess morphology and matched positive-control tissue before attributing the difference to fixation (standard IHC practice; HPA: High in cardiomyocytes). Keep section thickness, antibody incubation, and chromogen development consistent across the comparison so changes in signal can be interpreted more reliably (standard IHC practice).

What staining pattern should count as plausible ACADVL localisation?

Expect a granular cytoplasmic pattern in paraffin sections, consistent with the tissue staining profile and mitochondrial localisation of ACADVL (HPA: granular cytoplasmic profile; UniProt P49748: mitochondrion inner membrane). ACADVL has no annotated transmembrane segment, so a continuous cell-surface rim would warrant closer scrutiny (UniProt P49748: topology). Compare staining within identified cells against a heart-muscle positive control, where cardiomyocyte staining is high (HPA: High in cardiomyocytes; standard IHC practice). If a nuclear signal dominates, document it separately and check controls before calling it ACADVL, because additional nucleoplasmic and nucleolar localisation is uncertain in the subcellular dataset (HPA subcellular: uncertain additional locations).

Could isoforms or epitope accessibility explain discordant ACADVL staining?

ACADVL has 3 annotated isoforms, while the supplied antibody caption does not identify the recognised epitope (UniProt P49748: isoforms; caption PB10023). Check the antibody's epitope documentation before interpreting a negative section as absence of every isoform (standard IHC interpretation practice). The annotated mature chain spans residues 41–655, and the record includes modified residues such as acetyllysine at 51 and succinyllysine at 195 (UniProt P49748: processing and modified residues). Without an epitope map, these annotations cannot establish an effect on staining; compare positive-control tissue and retrieval conditions before proposing epitope-dependent loss (standard IHC practice).

How can IF help check an ambiguous chromogenic ACADVL pattern?

Use IF as a separate localisation check when granular cytoplasmic DAB staining is difficult to assign to individual cells (HPA: granular cytoplasmic profile; standard microscopy practice). Multiplex ACADVL with a validated cardiomyocyte marker in heart muscle, where cardiomyocytes show high tissue staining, and include single-label controls for each channel (HPA: High in cardiomyocytes; standard IF practice). Choose a far-red fluorophore if tissue autofluorescence obscures shorter-wavelength signal, and inspect unstained tissue before setting exposure (standard IF practice). Because ACADVL is associated with the mitochondrial inner membrane but its antibody epitope and membrane-facing side are unspecified, optimize permeabilisation empirically alongside mitochondrial-pattern controls (UniProt P49748: localisation and topology; caption PB10023; standard IF practice).

How do I reduce diffuse DAB staining without losing ACADVL signal?

First compare the stained section with a no-primary control to identify background from detection reagents or endogenous peroxidase (standard chromogenic IHC practice). Include a peroxidase-blocking step before HRP detection and keep DAB development time consistent across sections (standard chromogenic IHC practice). The selected image used 10% goat serum, 2 µg/ml primary antibody overnight at 4°C, and a peroxidase-conjugated secondary for 30 minutes at 37°C (caption PB10023). If background persists, assess blocking, washes, and primary-antibody concentration while preserving a granular cytoplasmic signal in a positive-control section (HPA: granular cytoplasmic profile; standard IHC optimization practice).

How should I score ACADVL staining across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION

Define the cell population and region of interest before scoring, then record the percentage of positive cells and staining intensity to calculate an H-score (standard IHC scoring practice). Score granular cytoplasmic signal separately from diffuse or nuclear staining, because the reported tissue pattern is granular cytoplasmic and additional nuclear localisation is uncertain (HPA: tissue profile; HPA subcellular: uncertain additional locations). For area-based analysis, report positive-cell density per mm² of viable tissue and normalise to the number or area of evaluable cells in the same region (standard image-analysis practice). Apply identical illumination, DAB thresholds, and counterstain criteria to all sections, with positive and no-primary controls reviewed alongside them (standard IHC quantification practice).

How can I distinguish true ACADVL staining from tissue artefact?

Give greatest weight to granular cytoplasmic staining in intact cells, consistent with the tissue profile and mitochondrial inner-membrane localisation (HPA: granular cytoplasmic profile; UniProt P49748: localisation). Check cell identity: cardiomyocytes show high staining, whereas chondrocytes were not detected in the supplied tissue survey (HPA: High in cardiomyocytes; HPA: Not detected in chondrocytes). Treat section-edge enhancement, staining over necrotic areas, and signal reproduced in a no-primary control as possible processing or detection artefacts (standard IHC interpretation practice). If staining appears chiefly nuclear, retain that observation but seek independent corroboration, because the reported nucleoplasmic and nucleolar locations are uncertain (HPA subcellular: uncertain additional locations).

Boster reagents

Best ACADVL / Very long-chain acyl-CoA dehydrogenase, mitochondrial IHC Antibodies

PB10023 has paraffin-section IHC images from human liver cancer, human ovarian cancer and rat heart, plus IF data from U2OS cells (catalog: image captions). M02607 lists human IHC (catalog: applications and reactivity).

Real IHC data

Anti-ACADVL Antibody ®

Cat # PB10023

View product →

M02607 is listed for human IHC, with no IHC image supplied (catalog: applications, reactivity and image captions). PB10023 has paraffin-section IHC images from human liver cancer, human ovarian cancer and rat heart, and an IF image from U2OS cells (catalog: image captions).

Which to pick: For tissue IHC, PB10023 has illustrated paraffin-section examples using EDTA retrieval at pH 8.0 and 2 μg/ml antibody; the fixative is unreported (PB10023 IHC captions). For IF/ICC, choose PB10023, which lists both applications and has an IF image from U2OS cells at 5 μg/ml (catalog: applications and PB10023 IF caption). For work across species, PB10023 lists human, mouse and rat reactivity, although its supplied IHC images cover only human and rat; M02607 is a rabbit monoclonal, clone 32A31, listed for human IHC (catalog: reactivity, applications, clone and image captions).

Each figure is that product's own IHC / IF validation image from its datasheet.

References

UniProt Consortium. UniProt entry P49748 (ACADV_HUMAN, Very long-chain acyl-CoA dehydrogenase, mitochondrial).
Human Protein Atlas. ACADVL tissue IHC expression (reliability: Enhanced).
Human Protein Atlas. ACADVL subcellular location (ICC-IF): Mainly localized to the mitochondria. In addition localized to the nucleoplasm and nucleoli..
Human Protein Atlas. ACADVL antibody validation summary (2 antibodies).
Proteomics Analysis Provides Insights into the Role of Lipid Metabolism in T2DM-Related Sarcopenia. ACS omega 2024 — PMC11308078.
AAV9 gene replacement therapy for respiratory insufficiency in very-long chain acyl-CoA dehydrogenase deficiency. Journal of inherited metabolic disease 2019 — PMC6739149.
Synthetic mRNA rescues very long-chain acyl-CoA dehydrogenase deficiency in patient fibroblasts and a murine model. Molecular genetics and metabolism 2023 — PMC9877169.
Unveiling the aging-immune axis in irritable bowel syndrome: a multi-omics and machine learning approach to biomarker discovery and validation. Frontiers in immunology 2026 — PMC13437292.
PubMed PMID:7668252 — UniProt-cited evidence.
PubMed PMID:8845838 — UniProt-cited evidence.
PubMed PMID:8554625 — UniProt-cited evidence.