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CERS5 / Ceramide synthase 5 · IHC design guide
Design Immunohistochemistry for CERS5
Plan chromogenic CERS5 IHC in paraffin sections with a human IHC-P–validated catalog antibody (datasheet: A08612-1). Interpret cytoplasmic and nucleolar tissue staining (HPA tissue IHC) alongside the ER membrane location (UniProt).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
CERS5 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 | IHC: cytoplasmic and nucleolar (HPA tissue IHC); ER membrane (UniProt) | |
| Staining pattern | Cytoplasmic and nucleolar staining in several tissues (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Testis+4 more · see all | |
| Negative control | Heart muscle+1 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A08612-1) | |
| Caveat | Nucleolar tissue staining differs from ER localisation (HPA tissue IHC; UniProt) | |
| Regulation | PAQR4 regulates CERS5 stability (UniProt) | |
| Isoform / epitope | 2 isoforms; epitope coverage across lumenal and cytoplasmic regions is unknown (UniProt) |
Section 1
Recommended CERS5 IHC & IF Protocols
Compare the catalog antibody’s IHC-P protocol with a published CERS5 staining method for gastric cancer tissue microarrays (PMC9552061).
| Sample | Tissue sections; selected-image fixative not specified (standard IHC workflow) |
| Fixation | Image fixative and duration unreported (datasheet A08612-1); 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-CERS5, 2.5 μg/mL (datasheet A08612-1) |
| 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 | CERS5-positive staining in sertoli cells of testis (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic and nucleolar expression in several tissues. No signal in the no-primary control. |
💡Decision noteStart with citrate pH 6.0 retrieval at 95–98 °C for 20 min (page antigen retrieval setting). The published method used microwave retrieval in water (PMC9552061).
Section 2
What Is the Expected CERS5 Staining Pattern?
CERS5 is a six-pass endoplasmic reticulum membrane protein (UniProt Q8N5B7 topology). In tissue IHC, expect cytoplasmic staining, sometimes with nucleolar staining, in several tissues (HPA: tissue profile). Sertoli cells provide a strong positive example; glandular cells in adrenal gland, appendix, breast, colon and duodenum show medium staining (HPA: tissue IHC). HPA rates the tissue profile Approved, with medium consistency between staining and RNA expression (HPA: reliability).
| Sertoli cells stain strongly; adjacent structures show less staining. | This fits the clearest listed tissue positive (HPA: High in testis Sertoli cells). Assess signal within the identified cells, rather than treating every stained area of the section as equally informative (general IHC practice). |
| Glandular cells show cytoplasmic staining, with possible nucleolar staining. | This matches the reported tissue profile (HPA: cytoplasmic and nucleolar expression). Medium glandular staining is listed in adrenal gland, appendix, breast, colon and duodenum (HPA: tissue IHC). Nucleolar signal alone should not replace the expected cellular context (HPA: tissue profile). |
| Staining is confined to an unexpected compartment, such as uniform nuclear fill without a cytoplasmic component. | Check for artefact before calling it CERS5: the reported tissue pattern includes cytoplasm and nucleoli, while UniProt places CERS5 in the endoplasmic reticulum membrane (HPA: tissue profile; UniProt Q8N5B7 subcellular location). A nucleolus and a nuclear membrane are distinct structures (general cell biology). |
| Cardiomyocytes or smooth muscle cells stain strongly, or staining spreads across unrelated cell types. | Treat this as a specificity warning, not proof of cross-reactivity: those cells are listed as not detected in their respective tissues (HPA: heart muscle and smooth muscle IHC). Cross-reactivity or endogenous detection activity can produce misleading chromogenic signal (general IHC practice). |
| A known-positive cell population has no visible signal. | An absent result in Sertoli cells conflicts with the listed high staining (HPA: testis IHC). Check tissue identity and the detection workflow before inferring that the sample lacks CERS5 (general IHC practice). HPA's Approved tissue rating still carries only medium staining–RNA consistency (HPA: reliability). |
💡Expected CERS5 appearanceA convincing positive shows high staining in testis Sertoli cells or medium staining in the listed glandular cells, with a cytoplasmic component and possible nucleolar signal (HPA: tissue IHC); strong signal limited to HPA-negative cell types is suspect (HPA: heart muscle and smooth muscle IHC).
| Membrane topology | CERS5 has 6 transmembrane segments in an endoplasmic reticulum membrane protein (UniProt Q8N5B7 topology). Its lumenal residues 1–46 and cytoplasmic residues 332–392 are on different sides of that membrane (UniProt Q8N5B7 topology). Without an epitope map, their relevance to this antibody's staining is unknown. |
| IF/ICC Q: Is nuclear rim staining expected? | Yes, as an IF/ICC observation: HPA reports the nuclear membrane as the main approved location, with additional endoplasmic reticulum and primary cilium locations (HPA: subcellular ICC-IF). This does not make a nuclear rim an IHC-P requirement; assess paraffin sections against the tissue IHC profile (HPA: tissue IHC). |
| Isoforms and processing | UniProt lists 2 isoforms and a single 1–392 chain, with no signal peptide or propeptide (UniProt Q8N5B7). The supplied record does not map the IHC antibody epitope to either isoform, so isoform-specific staining cannot be assigned. |
| Validation scope | HPA lists IHC as Approved for antibody HPA026589 and ICC as Approved for HPA006780 (HPA: antibody validation). These are application-specific entries; neither status alone establishes that both antibodies produce the same compartment pattern in paraffin sections. |
| Situation | Likely cause | Next action |
|---|---|---|
| No staining in testis Sertoli cells. | The positive population may have been misidentified, or the staining workflow may have failed (general IHC practice); Sertoli cells are listed as High (HPA: testis IHC). | Confirm the cell population and inspect an appropriate positive control. Check the recorded antibody, retrieval, dilution and detection steps against the validated IHC-P workflow (general IHC practice). No CERS5-specific retrieval condition or dilution is supplied here. |
| Only widespread pale color is visible. | Diffuse background can arise from nonspecific antibody binding or detection chemistry (general IHC practice). It does not reproduce the cell-selective tissue pattern (HPA: tissue IHC). | Review blocking, washes and antibody concentration; compare a control processed without primary antibody (general IHC practice). Score a cell population only when its signal is distinguishable from the background. |
| Strong chromogen appears in cardiomyocytes or smooth muscle cells. | These are listed as not detected in heart muscle and smooth muscle (HPA: tissue IHC). Cross-reactivity or endogenous detection activity is possible, but the image alone cannot distinguish them (general IHC practice). | Compare the same cells in a no-primary control and inspect the detection system's endogenous-activity control steps (general IHC practice). Recheck tissue and cell identity before interpreting the stain. |
| A section shows nuclear fill but little cytoplasmic signal. | Uniform nuclear fill does not match the reported cytoplasmic and nucleolar tissue profile (HPA: tissue IHC). IF/ICC nuclear membrane staining describes a rim, not nuclear fill (HPA: subcellular ICC-IF). | Inspect unstained or control sections for pigment and background, and compare signal with cell boundaries and nucleoli (general IHC practice). Do not transfer the IF/ICC localization call directly to IHC-P. |
| Glandular staining is weaker than staining in the testis example. | The listed glandular populations are Medium, while testis Sertoli cells are High (HPA: tissue IHC). A weaker glandular result can therefore fit the reported distribution. | Identify and score the relevant cell type in each section before comparing intensity (general IHC practice). Investigate a complete absence of signal separately from a moderate result. |
| Nucleolar staining appears without convincing cytoplasmic staining. | HPA describes both cytoplasmic and nucleolar expression across several tissues (HPA: tissue profile), while UniProt places CERS5 at the endoplasmic reticulum membrane (UniProt Q8N5B7 subcellular location). An isolated nucleolar result is ambiguous. | Re-examine the cytoplasm in the same cells and compare a no-primary control (general IHC practice). Report the observed compartment explicitly instead of calling isolated nucleolar signal a definitive positive. |
Experiment evidence
IHC & IF images showing CERS5 staining
A08612-1 · IHCImmunohistochemistry of LASS5 in human brain tissue with LASS5 antibody at 2.5 μg/mL.
A08612-1 · IFImmunofluorescence of LASS5 in Human Brain cells with LASS5 antibody at 20 μg/mL.
A08612 · IFImmunofluorescence of LASS5 in human brain tissue with LASS5 antibody at 20 μg/mL.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000139624-CERS5/tissue/cerebral+cortex#imid_6509816
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000139624-CERS5/tissue/colon#imid_6513190
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000139624-CERS5/tissue/liver#imid_6513207
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000139624-CERS5/tissue/kidney#imid_6513192
HPA · ICC-IF SK-MEL-30Human Protein Atlas immunofluorescence, SK-MEL-30, antibody HPA006780; green = CERS5, blue = nucleus, red = microtubules.
HPA · ICC-IF SK-MEL-30Human Protein Atlas immunofluorescence, SK-MEL-30, antibody HPA006780; green = CERS5, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for CERS5 IHC & IF
🧪Run testis first and score Sertoli cells, where CERS5 staining is High (HPA: High in Sertoli cells). Use heart muscle as the negative tissue, with cardiomyocytes expected to show no detectable staining (HPA: Not detected in cardiomyocytes); on the testis slide, cells outside the scored Sertoli population should provide a background comparison, not an assumed biological negative.
Positive control tissue: Testis (Sertoli cells, HPA High)
Negative control tissue: Heart muscle (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show CERS5 in SK-MEL-30, U2OS, hTERT-RPE1, ASC52telo, RPTEC/TERT1, hTERT-RPE1 (serum starved), with annotated localisation: Nuclear membrane (approved) (HPA subcellular).
Technical controls: Include a no-primary (secondary-only) control, an isotype control matched to the primary antibody’s host species and clonality, and a CERS5 knockout specimen or cognate-peptide block if available. For chromogenic IHC, quench endogenous peroxidase and check whether any testis signal persists in the no-primary control (standard IHC practice).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected A08612-1 human brain IHC caption does not state a fixative (caption: fixative not stated); retrieval dependence is also unreported, so compare staining with and without antigen retrieval. The supplied evidence does not establish whether frozen sections or IF are easier, although intracellular CERS5 detection by IF requires permeabilization (UniProt Q8N5B7: endoplasmic reticulum membrane; HPA subcellular: nuclear membrane and endoplasmic reticulum). In testis, score Sertoli cells separately from neighboring cells to avoid a cell-composition artefact (HPA: High in Sertoli cells).
HPA tissue IHC evidence for CERS5
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — 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 |
|---|---|---|---|---|
| Testis | Sertoli cells | High | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | Medium | Protein (IHC) | HPA → |
| Appendix | Glandular cells | Medium | Protein (IHC) | HPA → |
| Breast | Glandular cells | Medium | Protein (IHC) | HPA → |
| Cerebral cortex | Glial cells | Medium | Protein (IHC) | HPA → |
Section 3
Advanced CERS5 IHC Tips
CERS5 is a six-pass endoplasmic reticulum membrane enzyme; compartment and cell-type controls are central to interpreting its IHC signal (UniProt Q8N5B7; HPA tissue IHC).
How should I retrieve CERS5 in paraffin sections when staining is weak?
Use citrate pH 6.0 heat-induced retrieval at 95–98 °C for 20 min as the starting condition for paraffin CERS5 IHC (page retrieval rule). Let sections cool in buffer, then compare staining with a matched section processed identically apart from retrieval; this helps distinguish poor epitope access from detection failure (standard IHC practice). CERS5 has 6 transmembrane segments, so epitope access can depend on processing and antibody specificity (UniProt Q8N5B7 topology; standard IHC practice). If staining remains weak, test a different retrieval buffer or pH on serial sections while holding antibody concentration and detection constant (standard IHC practice). Score the expected cellular pattern alongside background before accepting a stronger signal (HPA tissue IHC; standard IHC practice).
Could fixation be responsible for weak or uneven CERS5 staining?
Target-specific fixation sensitivity for CERS5 is unknown from the supplied evidence; the selected tissue-IHC caption does not state a fixative (A08612-1 caption). Record the fixative, fixation duration, section age, and processing history before comparing slides, because these variables can affect antigen access in paraffin IHC (standard IHC practice). Run serial sections through the same citrate pH 6.0 retrieval at 95–98 °C for 20 min, changing only one processing variable per comparison (page retrieval rule; standard IHC practice). Use a matched positive-control section and a primary-antibody omission control to separate lost staining from nonspecific detection (standard IHC practice). Do not infer fixation tolerance from the reported tissue pattern (HPA tissue IHC).
Which cellular compartments should count as plausible CERS5 staining?
Prioritize cytoplasmic staining compatible with an endoplasmic reticulum membrane protein, and evaluate perinuclear or nuclear-rim staining in context (UniProt Q8N5B7 subcellular location; HPA tissue IHC; HPA subcellular). HPA tissue IHC describes cytoplasmic and nucleolar expression, while its cell-imaging summary places CERS5 mainly at the nuclear membrane and additionally at the endoplasmic reticulum and primary cilium (HPA tissue IHC; HPA subcellular). These observations come from different assay contexts, so a nucleolar or ciliary signal alone should prompt antibody-specific validation in the stained section (HPA tissue IHC; HPA subcellular; standard IHC practice). Compare compartment patterns across cells and controls at the same magnification and detection exposure (standard IHC practice).
How could epitope position change the CERS5 IHC pattern?
CERS5 has 2 reported isoforms and 6 transmembrane segments; its N-terminal residues 1–46 are lumenal and residues 332–392 are cytoplasmic (UniProt Q8N5B7). Check the antibody's disclosed immunogen or mapped epitope against both isoforms before treating staining as total CERS5 (UniProt Q8N5B7; standard IHC practice). A reported glycosylation site at residue 26 makes the local N-terminal context relevant if the epitope lies there, but the supplied evidence does not establish an antibody-specific staining effect (UniProt Q8N5B7). Compare serial sections with appropriate positive and omission controls, keeping retrieval and detection identical, when testing whether a distinct pattern is reproducible (standard IHC practice).
How should I check a CERS5 pattern by multiplex IF?
For the separate IF/ICC workflow, pair CERS5 with a marker for the cell population being evaluated, such as a glial-cell marker when examining cerebral cortex, and check whether the cellular assignments agree (HPA tissue IHC; standard IF practice). Choose fluorophores after measuring tissue autofluorescence in an unstained section; use a spectrally separated channel and matched single-label controls to assess bleed-through (standard IF practice). If the antibody epitope is cytoplasmic, permeabilise to expose that side of the membrane; a lumenal epitope may require different access, so verify the disclosed epitope before tuning detergent treatment (UniProt Q8N5B7 topology; standard IF practice). HPA reports nuclear-membrane and endoplasmic-reticulum localisation in cell imaging, which provides a pattern to compare, not an IF protocol for this antibody (HPA subcellular).
What should I change when CERS5 IHC has diffuse brown background?
First inspect a section processed without primary antibody to assess detection-system staining, and compare it with a matched stained section (standard IHC practice). Apply a peroxidase block before chromogenic development, control nonspecific binding with an appropriate protein block, and shorten DAB development if the entire section darkens (standard IHC practice). The selected caption reports CERS5 tissue IHC at 2.5 µg/mL in human brain, but it gives no fixative or detection details; treat that concentration as image context rather than a universal starting dilution (A08612-1 caption). Titrate the catalog antibody and detection time on serial sections while retaining the same citrate pH 6.0 retrieval (page retrieval rule; standard IHC practice).
How should I quantify CERS5 across differently composed tissue regions? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and region before scoring, then report the percentage of positive cells and an intensity-based H-score on the same segmentation rules for every section (standard IHC practice). If cell numbers vary substantially, report positive-cell density per mm² alongside the fraction of eligible cells that stain (standard IHC practice). Normalize comparisons to the number or area of the relevant cells, staining batch, and matched control-section response rather than whole-section brown area alone (standard IHC practice). HPA reports high staining in testis Sertoli cells and medium staining in cerebral-cortex glial cells, so preserve cell-type labels when comparing those patterns (HPA tissue IHC). Keep retrieval, antibody concentration, and DAB development matched across the quantified set (standard IHC practice).
When is a CERS5-positive IHC signal likely to be artefactual?
A reproducible cellular signal with cytoplasmic or perinuclear distribution is compatible with reported CERS5 localisation, but compartment agreement alone does not establish antibody specificity (UniProt Q8N5B7; HPA tissue IHC; HPA subcellular; standard IHC practice). Scrutinize strong staining restricted to section edges, folds, or necrotic areas, and staining reproduced by primary-antibody omission, as possible processing or detection artefacts (standard IHC practice). Check cell identity: HPA reports medium staining in cerebral-cortex glia but no detected staining in cardiomyocytes or smooth-muscle cells (HPA tissue IHC). Endogenous peroxidase can mimic DAB positivity, so interpret isolated brown deposits against the peroxidase-blocked and omission controls (standard IHC practice). HPA calls its tissue-IHC reliability Approved with medium staining–RNA consistency; treat borderline patterns cautiously (HPA tissue IHC).
Boster reagents
Best CERS5 / Ceramide synthase 5 IHC Antibodies
A08612-1 has human brain tissue IHC data (catalog image: human brain tissue) and human brain cell IF data (catalog image: human brain cells); A08612 has human brain tissue IF data (catalog image: human brain tissue).
Real IHC data
Anti-LASS5 CERS5 Antibody
Cat # A08612-1
Real IF data
Anti-LASS5 CERS5 Antibody
Cat # A08612
A08612-1 supports IHC-P and IF (catalog: applications), with IHC shown in human brain tissue at 2.5 μg/mL and IF shown in human brain cells at 20 μg/mL (catalog image captions). A08612 supports IF (catalog: applications), shown in human brain tissue at 20 μg/mL (catalog image caption).
Which to pick: Choose A08612-1 for paraffin-section IHC (catalog: IHC-P); its own human brain tissue IHC image uses 2.5 μg/mL (catalog image caption), and the fixative is unreported (catalog image caption). For IF, consider either SKU (catalog: applications); the images show human brain cells for A08612-1 and human brain tissue for A08612 (catalog image captions), while ICC validation is unreported (catalog: applications). Both list Human, Mouse and Rat reactivity (catalog: reactivity), but the supplied images establish only human-sample data (catalog image captions); clonality is unreported for both (catalog: clone).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q8N5B7 (CERS5_HUMAN, Ceramide synthase 5).
- Human Protein Atlas. CERS5 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. CERS5 subcellular location (ICC-IF): Mainly localized to the nuclear membrane. In addition localized to the endoplasmic reticulum and primary cilium..
- Human Protein Atlas. CERS5 antibody validation summary (2 antibodies).
- High expression of ceramide synthase 5 predicts a poor prognosis in gastric cancer. Translational cancer research 2022 — PMC9552061.
- High CerS5 expression levels associate with reduced patient survival and transition from apoptotic to autophagy signalling pathways in colorectal cancer. The journal of pathology. Clinical research 2015 — PMC4858121.
- Ceramide Synthase 5 Deficiency Aggravates Dextran Sodium Sulfate-Induced Colitis and Colon Carcinogenesis and Impairs T-Cell Activation. Cancers 2020 — PMC7409364.
- Comprehensive profiling of lipid metabolic reprogramming expands precision medicine for HCC. Hepatology (Baltimore, Md.) 2025 — PMC11902616.
- PubMed PMID:14702039 — UniProt-cited evidence.
- PubMed PMID:16541075 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for CERS5 (Ceramide synthase 5). Expected localisation IHC: cytoplasmic and nucleolar (HPA tissue IHC); ER membrane (UniProt). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: A08612-1 at 2.5 μg/mL (the product's own IHC validation image is shown). Controls (HPA): positive Testis (Sertoli cells); negative Heart muscle. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Tissue sections; selected-image fixative not specified (standard IHC workflow); 2. Fixation: Image fixative and duration unreported (datasheet A08612-1); 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-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-CERS5, 2.5 μg/mL (datasheet A08612-1); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A08612-1); Caveat: Nucleolar tissue staining differs from ER localisation (HPA tissue IHC; UniProt); Isoform / epitope: 2 isoforms; epitope coverage across lumenal and cytoplasmic regions is unknown (UniProt); Regulation: PAQR4 regulates CERS5 stability (UniProt). Evidence: UniProt Q8N5B7, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the nuclear membrane. In addition localized to…, Datasheet A08612-1, PMC9552061. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/CERS5/immunohistochemistry-ihc-guide.