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ACER3 / Alkaline ceramidase 3 · IHC design guide
Design Immunohistochemistry for ACER3
Use high-staining placenta decidual cells or kidney glomerular cells to plan ACER3 paraffin-section controls (HPA tissue IHC). This guide covers fixation, detection and interpretation using the catalog antibody’s IHC dilution of 1:100–1:300 (datasheet).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
ACER3 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 | ER and Golgi membranes expected at the molecular level (UniProt) | |
| Staining pattern | Endothelial cell staining; subcellular pattern unspecified (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Endometrium+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 | Antibody staining has low consistency with RNA expression (HPA tissue IHC) | |
| Regulation | No defined staining regulator (UniProt) | |
| Isoform / epitope | 2 isoforms; epitope coverage is unspecified (UniProt) |
Section 1
Recommended ACER3 IHC & IF Protocols
The catalog antibody’s IHC-P protocol is paired with a published mouse liver IHC workflow (PMC11873283).
| Sample | Paraffin-embedded human placenta tissue; fixative not specified (datasheet A10457-2) |
| Fixation | Image fixative and duration unreported (datasheet A10457-2); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat-induced epitope retrieval in citrate buffer, pH 6.0, 20 min at 95–98 °C (standard rule: cytoplasmic / membrane antigen) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% normal serum of the secondary host, 30 min, room temperature (standard) |
| Primary antibody | Rabbit anti-ACER3, 1:100-1:300 (datasheet A10457-2) |
| Primary incubation | Overnight at 4 °C (standard) |
| Detection | HRP-polymer secondary, DAB chromogen 5–10 min (standard) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | ACER3-positive staining in glandular cells of endometrium (HPA tissue IHC: High). HPA tissue profile: Expressed in endothelial cells. No signal in the no-primary control. |
💡Decision noteStart with citrate pH 6.0 retrieval at 95–98 °C for 20 min (page retrieval rule; UniProt Q9NUN7 membrane topology).
Section 2
What Is the Expected ACER3 Staining Pattern?
ACER3 is a 7-transmembrane protein of the endoplasmic reticulum and Golgi membranes (UniProt Q9NUN7 topology and subcellular location). In paraffin-section IHC, assess staining within the cell types HPA reports, including endometrial glandular cells, lung macrophages and placental decidual cells (HPA: High). HPA rates tissue IHC Approved, while reporting low consistency between antibody staining and RNA expression (HPA: reliability description).
| Staining is concentrated within cells, with a cytoplasmic or perinuclear appearance in a reported positive cell population. | This is compatible with ACER3 at ER and Golgi membranes (UniProt Q9NUN7 subcellular location). Resolve the positive cells by tissue morphology: HPA reports High staining in endometrial glandular cells, lung macrophages and placental decidual cells (HPA: tissue IHC). Chromogenic IHC alone does not prove which organelle contains the signal (general IHC interpretation). |
| Signal appears predominantly nuclear or outlines the cell surface, without a plausible intracellular component. | Reassess specificity: nuclear and surface-only patterns do not match the reported ER and Golgi membrane locations (UniProt Q9NUN7 subcellular location). Compare with a reported positive cell population and appropriate controls before calling the signal ACER3 (general IHC practice). |
| Strong staining appears in a cell population HPA lists as not detected, such as adipocytes in adipose tissue. | This conflicts with that specific HPA observation (HPA: Not detected in adipose-tissue adipocytes). Consider cross-reactivity or endogenous detection activity, and inspect a known-positive tissue on the same run (general IHC practice). HPA staining varies by cell population, so do not treat the entire tissue as a uniform negative (HPA: tissue IHC). |
| Color spreads across tissue or appears similarly in many unrelated structures, obscuring cell boundaries. | Treat this as background until controls show otherwise (general IHC practice). Check blocking, detection reagents, wash quality and counterstain, then score only signal that can be assigned to identifiable cells (general IHC practice). Broad staining alone is weak evidence because HPA reports low tissue RNA specificity but low staining–RNA consistency (HPA: tissue IHC and reliability description). |
| No convincing signal appears in a section containing an expected positive population. | First verify that the relevant cells are present: HPA reports High staining in kidney glomerular cells, skin fibroblasts and testis Sertoli cells, among others (HPA: tissue IHC). Check the run controls and IHC reagent performance (general IHC practice). A blank section does not establish absent ACER3 expression, particularly given HPA's reported staining–RNA inconsistency (HPA: reliability description). |
💡Expected ACER3 appearanceCall a positive result when identifiable HPA-reported cells show clear intracellular staining compatible with ER/Golgi membranes, preferably in a High HPA population; isolated nuclear, surface-only or diffuse tissue-wide color warrants specificity checks (UniProt Q9NUN7 subcellular location; HPA: tissue IHC; general IHC interpretation).
| Intracellular topology | ACER3 has 7 transmembrane segments and both cytoplasmic and lumenal regions (UniProt Q9NUN7 topology). The supplied record does not locate the antibody epitope; do not infer which region the antibody detects or prescribe target-specific retrieval from topology alone. |
| Choice of comparison cells | HPA reports High staining in several distinct populations, Medium in appendix lymphoid tissue, and Not detected in selected others (HPA: tissue IHC). Judge cells within their tissue context; a negative adipocyte observation does not make all cells in that section negative (HPA: tissue IHC). |
| Antibody evidence | The listed rabbit polyclonal antibody HPA070087 has IHC status Approved and no supplied ICC status (HPA: antibody validation). HPA also reports low consistency between tissue antibody staining and RNA expression; interpret borderline patterns with controls (HPA: reliability description; general IHC practice). |
| Isoforms and processing | UniProt lists 2 isoforms and a chain spanning residues 1–267, with no annotated signal peptide, propeptide or glycosylation sites (UniProt Q9NUN7 isoforms, processing and glycosylation). The supplied evidence does not show isoform-specific staining or shedding; avoid assigning either from IHC appearance. |
| IF/ICC Q&A: Is the IHC pattern sufficient to predict IF/ICC staining? | No. HPA summarizes the subcellular location as Membrane but provides no main location or ICC/IF image-bearing cell lines, and the listed antibody has no supplied ICC status (HPA: subcellular and antibody records). ER/Golgi localization is a hypothesis to test in IF/ICC, not an observed ICC pattern (UniProt Q9NUN7 subcellular location). |
| Situation | Likely cause | Next action |
|---|---|---|
| No staining in a reported High population. | The expected cells may be absent from the section, or an IHC step may have failed (HPA: tissue IHC; general IHC practice). | Confirm cell identity and section integrity, then review positive-run controls, antigen retrieval, antibody incubation and chromogenic detection records (general IHC practice). The payload supplies no ACER3-specific retrieval condition. |
| Signal is confined to nuclei or cell borders. | That distribution is discordant with ER/Golgi membrane localization (UniProt Q9NUN7 subcellular location); nonspecific staining is possible (general IHC interpretation). | Inspect a reported positive cell population and negative detection controls, and require an intracellular component before assigning ACER3 (HPA: tissue IHC; general IHC practice). |
| Adipocytes stain strongly in adipose tissue. | HPA lists adipose-tissue adipocytes as Not detected; cross-reactivity or endogenous detection activity may explain a conflicting signal (HPA: tissue IHC; general IHC practice). | Compare cell-level staining with a reported High population, then check reagent-only and detection controls before interpreting the adipocyte signal (HPA: tissue IHC; general IHC practice). |
| Diffuse brown color obscures cell morphology. | Background from blocking, washing or detection can make localization uninterpretable (general IHC practice). | Review reagent-only controls, blocking, wash steps and detection exposure; score only resolved cellular staining after background is controlled (general IHC practice). |
| A tissue looks negative despite broad expression reported elsewhere. | ACER3 is described as ubiquitously expressed by UniProt, while HPA reports cell-specific Not detected results and low staining–RNA consistency (UniProt Q9NUN7 tissue specificity; HPA: tissue IHC and reliability description). | Identify the actual cell population being scored and compare it with that tissue's HPA observation; use a reported High population as a run comparator (HPA: tissue IHC; general IHC practice). |
| A proposed retrieval change seems to reverse the result. | No ACER3-specific fixation sensitivity or retrieval effect is supplied; the change alone cannot establish target identity (supplied UniProt Q9NUN7 and HPA records). | Document both conditions and compare morphology, background and run controls under each; describe retrieval effects as observations from the experiment (general IHC practice). |
Experiment evidence
IHC & IF images showing ACER3 staining
A10457-2 · IHCImmunohistochemistry analysis of paraffin-embedded human placenta tissue, using PHCA Antibody. The picture on the right is blocked with the
A10457-2 · IFImmunofluorescence analysis of MCF7 cells, using PHCA Antibody. The picture on the right is blocked with the synthesized peptide.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000078124-ACER3/tissue/cerebral+cortex#imid_20444811
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000078124-ACER3/tissue/colon#imid_20444729
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000078124-ACER3/tissue/liver#imid_20444746
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000078124-ACER3/tissue/kidney#imid_20444731
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for ACER3 IHC & IF
🧪Run placenta first: decidual cells should stain (HPA: High in placenta decidual cells; selected A10457-2 tissue-IHC caption: placenta). Use adipose tissue as the negative comparator (HPA: adipocytes Not detected); on the placenta slide, internal negative cells should show counterstain without specific chromogenic signal, but the supplied HPA rows do not identify a negative placental cell type.
Positive control tissue: Endometrium (Glandular cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA carries no ICC-IF cell line for ACER3; derive a cell-line control from the positive tissue's cell type (Glandular cells) and confirm it by RNA or western blot first.
Technical controls: Include no-primary (secondary-only) and host-species- and clonality-matched isotype controls (standard IHC practice); use peptide blocking as a biological specificity control (selected A10457-2 tissue-IHC caption: signal blocked with synthesized peptide). For placental chromogenic IHC, quench endogenous peroxidase and block endogenous biotin if using biotin-based detection (standard IHC practice).
⚠️Feasibility: A target-specific fixation window and fixation effect are unreported, and the selected A10457-2 paraffin-section caption does not state a fixative (selected A10457-2 tissue-IHC caption). Antigen-retrieval dependency is unreported; optimize retrieval empirically for the paraffin-section assay (standard IHC practice). The supplied evidence does not establish whether frozen sections or IF/ICC would be easier; endogenous peroxidase or biotin may complicate placental chromogenic detection (standard IHC practice).
HPA tissue IHC evidence for ACER3
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Low 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 |
|---|---|---|---|---|
| Endometrium | Glandular cells | High | Protein (IHC) | HPA → |
| Kidney | Cells in glomeruli | High | Protein (IHC) | HPA → |
| Lung | Macrophages | High | Protein (IHC) | HPA → |
| Placenta | Decidual cells | High | Protein (IHC) | HPA → |
| Skin | Fibroblasts | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Breast | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Caudate | Glial cells | Not detected | Protein (IHC) | HPA → |
| Cerebellum | Cells in granular layer | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced ACER3 IHC Tips
Troubleshoot ACER3 staining in paraffin sections by checking retrieval, cellular pattern, controls and scoring before interpreting chromogenic signal.
What retrieval should I try first when ACER3 staining is weak?
Start with citrate buffer at pH 6.0 for heat induced retrieval at 95–98 °C for 20 minutes (page retrieval rule: cytoplasmic or membrane antigen). Allow slides to cool in the retrieval buffer, then keep section thickness, antibody incubation and detection conditions consistent while comparing runs (standard IHC practice). ACER3 has 7 transmembrane segments and resides in endoplasmic reticulum and Golgi membranes (UniProt Q9NUN7 topology and subcellular location). If staining remains weak, compare a longer retrieval time on adjacent sections as a controlled fallback, checking morphology and background (standard IHC practice). Use the same placenta control across runs; the catalog antibody has a paraffin section image with peptide blocking (A10457-2 caption).
How should I assess whether fixation is limiting ACER3 detection?
Target specific fixation sensitivity is unknown because the supplied ACER3 evidence gives no fixation method or comparison (A10457-2 caption: fixative not stated). Record the actual fixative, fixation duration and processing history for each block, and compare sections processed together (standard IHC practice). If signal varies between blocks, stain an internal control and adjacent sections in the same run before attributing the difference to ACER3 biology (standard IHC practice). The catalog image establishes staining in paraffin embedded placenta tissue, but its caption does not establish formalin fixation (A10457-2 caption). Keep retrieval at citrate pH 6.0, 95–98 °C for 20 minutes during this comparison (page retrieval rule).
What cellular pattern is plausible for ACER3 in chromogenic IHC?
Evaluate ACER3 as intracellular, membrane associated staining consistent with endoplasmic reticulum and Golgi localization (UniProt Q9NUN7 subcellular location). Its 7 transmembrane segments make a purely nuclear pattern difficult to reconcile with the annotated protein topology (UniProt Q9NUN7 topology). Inspect positive cells at high magnification and distinguish granular or perinuclear cytoplasmic signal from pigment, precipitate and nonspecific edge staining (standard IHC practice). In placenta, identify the stained cell population before comparing intensity: high staining is reported in decidual cells (HPA: placenta, decidual cells, High). Treat a discordant compartment or cell population as a prompt to review controls and morphology, since HPA reports low consistency between staining and RNA (HPA reliability description).
How do isoforms and membrane topology affect epitope interpretation?
ACER3 has 2 annotated isoforms, but the supplied evidence does not map the catalog antibody epitope to either one (UniProt Q9NUN7 isoforms; A10457-2 caption). The protein crosses membranes 7 times, with both cytoplasmic and lumenal intervals, so epitope accessibility may depend on its location (UniProt Q9NUN7 topology; standard IHC practice). Review the antibody's documented immunogen sequence against both isoforms before claiming isoform specificity (standard IHC practice). The supplied record lists 0 glycosylation sites and 0 modified residues; neither observation identifies the antibody's binding site (UniProt Q9NUN7 annotations). Peptide blocking in the placenta image supports competition by that peptide, but does not resolve isoform recognition (A10457-2 caption).
How can I adapt the ACER3 cell pattern check to multiplex IF?
For multiplex IF, pair ACER3 with a marker that identifies the expected cell population, such as a validated decidual cell marker in placenta (HPA: placenta, decidual cells, High; standard IF practice). Choose spectrally separated fluorophores and assess unstained tissue autofluorescence before assigning dim signal, especially in broad emission channels (standard IF practice). ACER3 has cytoplasmic and lumenal intervals across 7 membrane spans, so select permeabilisation conditions after confirming which side contains the antibody epitope (UniProt Q9NUN7 topology; standard IF practice). Compare permeabilised and matched control sections, keeping acquisition settings constant when judging overlap (standard IF practice). The supplied subcellular record has no ICC/IF images, so do not treat this multiplex pattern as independently validated (HPA subcellular record).
How do I reduce diffuse or cell independent brown staining?
Run a no primary control and inspect whether the same brown deposit persists, then check the secondary reagent and chromogen steps (standard chromogenic IHC practice). Block endogenous peroxidase before DAB development and use an appropriate protein block for nonspecific antibody binding (standard chromogenic IHC practice). Titrate the catalog antibody and shorten chromogen development on adjacent sections if staining obscures cell borders (standard IHC practice). Compare stained areas with morphology because high ACER3 staining is reported in specific cell populations, including lung macrophages and placental decidual cells (HPA: lung macrophages, High; HPA: placenta decidual cells, High). The peptide blocked placenta image is a product control, but does not replace section level background controls (A10457-2 caption; standard IHC practice).
What should I measure when comparing ACER3 across sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, then use an H-score from staining intensity and percentage of positive cells (standard IHC quantification practice). Record the percentage positive separately so a change in cell abundance is distinguishable from a change in staining strength (standard IHC quantification practice). For focal populations, count positive cells per mm² of viable tissue and normalise to the number of eligible cells in that same area (standard IHC quantification practice). Keep retrieval, DAB development, section selection and threshold rules consistent across samples (standard IHC practice). Report cell identity alongside scores because HPA lists high staining in several different cell types and notes low staining to RNA consistency (HPA tissue profile and reliability description).
When is an apparent ACER3 positive result unconvincing?
Treat a nuclear only signal cautiously because ACER3 is annotated in endoplasmic reticulum and Golgi membranes, with 7 transmembrane segments (UniProt Q9NUN7 subcellular location and topology). Verify that positive cells match the tissue context: HPA reports high staining in placental decidual cells and lung macrophages (HPA tissue profile). Staining confined to section edges, necrotic regions or the no primary control suggests artefact rather than convincing target localization (standard IHC practice). Persistent DAB signal after omitting the primary antibody warrants review of endogenous peroxidase blocking (standard chromogenic IHC practice). Interpret modest differences cautiously because the HPA tissue entry reports low consistency between antibody staining and RNA expression (HPA reliability description).
Boster reagents
Best ACER3 / Alkaline ceramidase 3 IHC Antibodies
The catalog lists A10457-2 for human and mouse IHC/IF (catalog: applications and reactivity), with images of paraffin-embedded human placenta and MCF7 cells (IHC/IF image captions).
Real IHC data
Anti-APHC ACER3 Antibody
Cat # A10457-2
A10457-2 is listed for IHC and IF in human and mouse samples (catalog: applications and reactivity). Its IHC image shows paraffin-embedded human placenta, and its IF image shows MCF7 cells (IHC/IF image captions).
Which to pick: For tissue IHC, choose A10457-2 for paraffin sections based on its human placenta image; the fixative is unreported (IHC image caption). For IF/ICC, A10457-2 has an IF image in MCF7 cells (IF image caption); its host is rabbit and clonality is unreported (catalog: host and clone). A10457-2 lists human and mouse reactivity (catalog: reactivity), but both image captions call the reagent “PHCA Antibody,” so confirm its identity before treating those images as ACER3 validation (IHC/IF image captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q9NUN7 (ACER3_HUMAN, Alkaline ceramidase 3).
- Human Protein Atlas. ACER3 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. ACER3 subcellular location (ICC-IF): Membrane.
- Human Protein Atlas. ACER3 antibody validation summary (1 antibodies).
- Targeting the ceramidase ACER3 attenuates cholestasis in mice by mitigating bile acid overload via unsaturated ceramide-mediated LXRβ signaling transduction. Nature communications 2025 — PMC11873283.
- Circular RNA circ_0001955 promotes hepatocellular carcinoma tumorigenesis by up-regulating alkaline ceramidase 3 expression through microRNA-655-3p. Bioengineered 2022 — PMC8973869.
- PubMed PMID:11356846 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
- PubMed PMID:16554811 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for ACER3 (Alkaline ceramidase 3). Expected localisation ER and Golgi membranes expected at the molecular level (UniProt). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: A10457-2 at 1:100-1:300 (the product's own IHC validation image is shown). Controls (HPA): positive Endometrium (Glandular cells); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human placenta tissue; fixative not specified (datasheet A10457-2); 2. Fixation: Image fixative and duration unreported (datasheet A10457-2); verify before use.; 3. Sectioning: 4–5 µm sections on charged slides (standard); 4. Deparaffinisation: Xylene, graded ethanol series to water (standard); 5. Antigen retrieval: Heat-induced epitope retrieval in citrate buffer, pH 6.0 , 20 min at 95–98 °C…; 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% normal serum of the secondary host, 30 min , room temperature (standard); 8. Primary antibody: Rabbit anti-ACER3, 1:100-1:300 (datasheet A10457-2); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across paraffin sections. (standard IHC practice; not target-specific); Caveat: Antibody staining has low consistency with RNA expression (HPA tissue IHC); Isoform / epitope: 2 isoforms; epitope coverage is unspecified (UniProt); Regulation: No defined staining regulator (UniProt). Evidence: UniProt Q9NUN7, HPA tissue IHC · Approved, HPA ICC-IF · Membrane, Datasheet A10457-2, PMC11873283. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/ACER3/immunohistochemistry-ihc-guide.