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CERS6 / Ceramide synthase 6 · IHC design guide
Design Immunohistochemistry for CERS6
Plan CERS6 IHC on paraffin sections using the reported nuclear tissue pattern (HPA tissue IHC) and the ER membrane annotation (UniProt). The guide helps select tissue controls and interpret staining in light of low antibody–RNA concordance (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
CERS6 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 | Nuclear in tissues (HPA tissue IHC); ER membrane (UniProt) | |
| Staining pattern | Nuclear 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 | Cerebellum+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 sections. (standard IHC practice; not target-specific) | |
| Caveat | Antibody staining has low concordance with RNA (HPA tissue IHC) | |
| Regulation | Staining induction unreported (UniProt) | |
| Isoform / epitope | 2 isoforms; epitope coverage needs checking (UniProt) |
Section 1
Recommended CERS6 IHC & IF Protocols
The catalog antibody’s IHC-P protocol is accompanied by published CERS6 staining methods for ESCC tissue arrays (PMC12594893) and mouse leukemia xenografts (PMC4695011).
| Sample | Tissue sections; selected-image fixative not specified (standard IHC workflow) |
| Fixation | Image fixative and duration unreported (datasheet A07033); 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-CERS6, 2.5 μg/mL (datasheet A07033) |
| 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 | CERS6-positive staining in cells in granular layer of cerebellum (HPA tissue IHC: High). HPA tissue profile: Nuclear expression in several tissues. No signal in the no-primary control. |
💡Decision noteStart with citrate pH 6.0 HIER at 95–98 °C for 20 min (page antigen retrieval); the xenograft study used pH 8.4 retrieval (PMC4695011).
Section 2
What Is the Expected CERS6 Staining Pattern?
CERS6 is a seven-pass endoplasmic reticulum (ER) membrane protein, so target-consistent staining would be intracellular and ER-associated (UniProt Q6ZMG9 topology). For tissue selection, HPA reports high staining in cerebellar granular-layer cells and medium staining in several other cell types (HPA tissue IHC). HPA also describes nuclear expression; its IHC reliability is Approved, with low consistency between antibody staining and RNA expression (HPA tissue IHC).
| Granular-layer cells stain strongly; signal appears cytoplasmic or perinuclear (HPA: High in cerebellar granular-layer cells; UniProt Q6ZMG9: ER membrane). | The cell type and ER-associated distribution fit the strongest supplied tissue observation and the annotated location. Use the cerebellar section to judge whether staining is cellular and patterned, while retaining the HPA reliability caveat (HPA tissue IHC: Approved; low staining–RNA consistency). |
| Staining is exclusively nuclear, including in granular-layer cells (HPA tissue IHC: nuclear expression in several tissues). | This matches HPA’s reported IHC profile but conflicts with the UniProt ER-membrane annotation (HPA tissue IHC; UniProt Q6ZMG9). Treat nuclear-only staining as unresolved; examine cellular distribution and controls before assigning it to CERS6. Nuclear location alone cannot establish specificity. |
| Strong staining appears in adipocytes or cardiomyocytes (HPA tissue IHC: Not detected in these cells). | This differs from the supplied cell-specific IHC observations and raises concern for cross-reactivity or endogenous chromogenic activity (HPA tissue IHC; general IHC practice). Check cell identity, compare the no-primary control, and interpret only staining attributable to the primary antibody. |
| Color spreads across stroma or tissue without clear cellular boundaries (general IHC practice). | A diffuse deposit is difficult to assign to the ER or to HPA-reported positive cells (UniProt Q6ZMG9; HPA tissue IHC). Assess the no-primary control and review blocking, washes, detection conditions, and counterstain as general chromogenic IHC checks. |
| Granular-layer cells show no detectable signal (HPA: High in cerebellar granular-layer cells). | The expected positive tissue has failed to reproduce the supplied HPA observation; this does not by itself prove CERS6 is absent (HPA tissue IHC; general IHC practice). Review the antibody’s validated IHC-P conditions, detection reagents, and tissue integrity before interpreting a negative result. |
💡Expected CERS6 appearanceA credible positive is strong, cellular staining in cerebellar granular-layer cells with an ER-compatible intracellular distribution; diffuse deposits or staining in HPA-negative cell types warrant investigation, while HPA-reported nuclear staining remains a localization discrepancy (HPA tissue IHC; UniProt Q6ZMG9).
| Membrane topology (UniProt Q6ZMG9 topology) | CERS6 has 7 transmembrane segments with lumenal and cytoplasmic regions (UniProt Q6ZMG9). Epitope location could affect antibody access, but the supplied record does not identify the catalog antibody’s epitope or establish a CERS6-specific antigen-retrieval requirement. |
| IHC validation (HPA antibodies; HPA tissue IHC) | HPA044683 is listed as IHC Approved; the tissue summary reports low consistency between staining and RNA expression (HPA antibodies; HPA tissue IHC). Approved status supports use of the reported pattern for comparison but does not resolve its nuclear-versus-ER discrepancy. |
| Cell-specific tissue contrast (HPA tissue IHC) | Cerebellar granular-layer cells are High; bone-marrow hematopoietic cells, cortical neurons, hippocampal glia, fallopian-tube glandular cells, and smooth-muscle cells are Medium (HPA tissue IHC). Adipocytes and cardiomyocytes are Not detected, providing cell-specific comparisons within the supplied observations. |
| Isoforms and epitope coverage (UniProt Q6ZMG9) | UniProt lists 2 isoforms (UniProt Q6ZMG9). The payload gives no antibody epitope or isoform coverage, so differing staining cannot be assigned to one isoform. The listed glycosylation site at residue 18 likewise does not establish an effect on IHC staining. |
| IF evidence boundary (HPA subcellular; HPA antibodies) | HPA supplies no CERS6 ICC-IF image, main subcellular location, or ICC validation entry for HPA044683 (HPA subcellular; HPA antibodies). Its ICC-IF record therefore cannot independently confirm the compartment seen in paraffin-section IHC. |
| Situation | Likely cause | Next action |
|---|---|---|
| Nuclear-only signal dominates the IHC section (HPA tissue IHC: nuclear expression). | HPA’s reported nuclear pattern conflicts with the ER-membrane annotation; the supplied evidence does not identify the reason (HPA tissue IHC; UniProt Q6ZMG9). | Record nuclear and cytoplasmic staining separately, inspect the no-primary control, and avoid scoring nuclear signal as confirmed ER-localized CERS6 (general IHC practice; UniProt Q6ZMG9). |
| Cerebellar granular-layer cells are blank (HPA: High in these cells). | A failed stain or detection step is possible; absence of CERS6 cannot be inferred from one blank section (HPA tissue IHC; general IHC practice). | Confirm tissue integrity and primary-antibody application, then check the catalog antibody’s IHC-P instructions and the chromogenic detection run (general IHC practice). |
| Diffuse color obscures cell boundaries (general IHC practice). | Background from detection reagents, incomplete blocking, or insufficient washing is possible in chromogenic IHC (general IHC practice). | Compare a no-primary control; review blocking, washes, and detection conditions before judging cellular staining (general IHC practice). |
| HPA-negative cells stain strongly (HPA: adipocytes and cardiomyocytes Not detected). | Cross-reactivity or endogenous detection activity is possible, especially if staining persists without primary antibody (HPA tissue IHC; general IHC practice). | Inspect the no-primary control and compare the same cell type across sections; report the discrepancy rather than treating it as established CERS6 expression (general IHC practice; HPA tissue IHC). |
| Medium-level tissues vary across sections (HPA: Medium in several listed cell types). | Section quality or staining-run variation may affect the comparison; HPA also reports low staining–RNA consistency (general IHC practice; HPA tissue IHC). | Compare equivalent cell types and a cerebellar positive section within the run; score cell type, compartment, and intensity separately (HPA tissue IHC; general IHC practice). |
| Can IF/ICC settle the nuclear-versus-ER question? (HPA tissue IHC; UniProt Q6ZMG9). | The supplied HPA ICC-IF record has no images or assigned main location, and HPA044683 has no ICC validation entry (HPA subcellular; HPA antibodies). | Treat IF/ICC as a separate validation question; this IHC-P evidence provides no supported IF/ICC protocol or confirmed IF localization (HPA subcellular; HPA antibodies). |
Experiment evidence
IHC & IF images showing CERS6 staining
A07033 · IHCImmunohistochemistry of LASS6 in mouse brain tissue with LASS6 antibody at 2.5 μg/mL.
A07033 · IFImmunofluorescence of LASS6 in Mouse Brain cells with LASS6 antibody at 5 μg/mL.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000172292-CERS6/tissue/cerebral+cortex#imid_12083179
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000172292-CERS6/tissue/colon#imid_12083113
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000172292-CERS6/tissue/liver#imid_12083114
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000172292-CERS6/tissue/kidney#imid_12083115
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for CERS6 IHC & IF
🧪Run cerebellum first and score cells in the granular layer, where CERS6 staining is High (HPA: cerebellum, cells in granular layer, High). Use adipose tissue as the negative comparison and score adipocytes, where staining is Not detected; on the cerebellar slide, treat cells showing counterstain alone as internal negative comparators without assuming that a particular neighboring cell type lacks CERS6 (HPA: adipose tissue, adipocytes, Not detected; standard IHC practice).
Positive control tissue: Cerebellum (Cells in granular layer, 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 CERS6; derive a cell-line control from the positive tissue's cell type (Cells in granular layer) and confirm it by RNA or western blot first.
Technical controls: Include no-primary and secondary-only controls, plus a host-species- and immunoglobulin-class-matched isotype control for a monoclonal antibody or matched nonimmune IgG for a polyclonal antibody; use knockout tissue, if available, or immunizing-peptide blocking, if the peptide is available, to test specificity (standard IHC practice). Block endogenous peroxidase for chromogenic detection and assess cerebellar pigment as a possible staining artefact (standard IHC practice).
⚠️Feasibility: A CERS6-specific fixation window or fixation effect is unreported, and the selected A07033 mouse-brain IHC caption does not state a fixative (A07033 caption). Antigen-retrieval dependence is unreported, so optimize retrieval on the positive and negative tissues before interpreting staining (supplied evidence; standard IHC practice). The supplied evidence does not establish that frozen-section IHC or IF/ICC is easier than paraffin IHC; HPA lists no ICC-IF image-bearing cell lines, and brain pigment warrants inspection during chromogenic scoring (HPA subcellular; standard IHC practice).
HPA tissue IHC evidence for CERS6
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 |
|---|---|---|---|---|
| Cerebellum | Cells in granular layer | High | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | Medium | Protein (IHC) | HPA → |
| Cerebral cortex | Neuronal cells | Medium | Protein (IHC) | HPA → |
| Fallopian tube | Glandular cells | Medium | Protein (IHC) | HPA → |
| Hippocampus | Glial cells | Medium | 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 → |
| Epididymis | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Heart muscle | Cardiomyocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced CERS6 IHC Tips
Troubleshoot CERS6 staining in paraffin section IHC using the page retrieval method, the catalog antibody’s tissue image, and compartment aware controls.
Which retrieval conditions should I start with for CERS6 in paraffin sections?
Start with citrate buffer at pH 6.0, heated to 95–98 °C for 20 min (page CERS6 IHC retrieval setting). Allow sections to cool in the retrieval buffer, then compare stained sections with a no primary control processed through the same chromogenic workflow (standard IHC practice). If staining is weak, adjust heating time in small increments while holding antibody dilution and detection conditions constant (standard IHC practice). Excessive heating can damage section morphology and make granular cytoplasmic staining harder to judge (standard IHC practice). Record retrieval conditions for each run because CERS6 has 7 membrane spanning segments and its epitope is unspecified (UniProt Q6ZMG9 topology; supplied antibody evidence).
How should I handle fixation when CERS6 staining is weak or uneven?
Target specific fixation sensitivity is unknown because the supplied CERS6 tissue image caption does not state its fixative (A07033 tissue IHC caption). For paraffin sections, document fixation duration and processing history, and compare sections prepared under matched conditions before changing antibody concentration (standard IHC practice). Uneven staining across a section warrants inspection for folds, drying and incomplete reagent coverage (standard IHC practice). Keep retrieval at citrate pH 6.0, 95–98 °C for 20 min while assessing fixation related variation (page CERS6 IHC retrieval setting). Do not infer a CERS6 specific fixation effect from tissue staining patterns or membrane topology (HPA tissue IHC profile; UniProt Q6ZMG9 topology).
What staining compartment is plausible for CERS6, and how should I assess nuclear signal?
CERS6 is annotated at the endoplasmic reticulum membrane, making a cytoplasmic, often perinuclear distribution plausible in chromogenic sections (UniProt Q6ZMG9 subcellular location; standard IHC interpretation). The HPA tissue profile instead reports nuclear expression in several tissues, with approved reliability but low consistency between staining and RNA data (HPA tissue IHC). Assess nuclear signal separately from cytoplasmic signal rather than combining them into one positive score (standard IHC practice). Compare the compartments across cells in the same section, and inspect a no primary control for nuclear chromogen deposition (standard IHC practice). HPA supplies no ICC/IF images that independently resolve this localisation discrepancy (HPA subcellular record).
How does an unknown epitope affect interpretation of CERS6 IHC?
CERS6 has 2 annotated isoforms, while the supplied antibody evidence does not identify an epitope or establish isoform coverage (UniProt Q6ZMG9 isoforms; supplied antibody evidence). Its 7 membrane spanning segments separate lumenal and cytoplasmic regions, so antibody access can depend on where the epitope lies (UniProt Q6ZMG9 topology; standard IHC practice). Compare citrate pH 6.0 retrieval with an unretrieved section while keeping detection settings matched (page CERS6 IHC retrieval setting; standard IHC practice). A changed staining pattern alone cannot identify the recognized isoform or place the epitope on one membrane side (standard IHC interpretation). Obtain epitope mapping or independent validation before making either claim (standard antibody validation practice).
How can I assess CERS6 by IF alongside a cell identity marker?
Treat IF as a separate assay: the supplied CERS6 image documents tissue IHC in mouse brain at 2.5 µg/mL, with no IF validation or fixation condition stated (A07033 tissue IHC caption). Multiplex CERS6 with a validated marker for the cell population being examined and use single stain controls to check channel overlap (standard IF practice). Choose a fluorophore channel with low tissue autofluorescence and inspect an unstained section before assigning weak signal to CERS6 (standard IF practice). Because the epitope is unspecified, optimise mild permeabilisation against an IF control; access requirements differ for lumenal and cytoplasmic membrane faces (UniProt Q6ZMG9 topology; standard IF practice). Interpret compartment agreement with endoplasmic reticulum localisation cautiously (UniProt Q6ZMG9 subcellular location).
How can I separate CERS6 signal from chromogenic background?
Run no primary and detection only controls to identify chromogen deposited without the CERS6 antibody (standard chromogenic IHC practice). Include a peroxidase block before enzyme based detection and check whether residual colour appears in the controls (standard chromogenic IHC practice). Optimise blocking and antibody concentration one variable at a time; the selected tissue IHC image used 2.5 µg/mL in mouse brain (A07033 tissue IHC caption; standard IHC practice). Diffuse colour over the entire section, tissue edges or damaged areas should be scored separately from cellular staining (standard IHC interpretation). Judge any remaining signal against the expected endoplasmic reticulum membrane location and the HPA nuclear pattern discrepancy (UniProt Q6ZMG9 subcellular location; HPA tissue IHC).
How should I score CERS6 staining across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, then record percentage positive cells and staining intensity as a compartment specific H-score (standard IHC scoring practice). For a spatial question, report positive cell density per mm² of viable tissue and exclude folds, section edges and necrotic regions (standard IHC quantification practice). Normalise counts to the number of eligible cells or measured viable area, and keep exposure, threshold and counterstain settings consistent (standard IHC quantification practice). Report nuclear and cytoplasmic results separately because UniProt places CERS6 at the endoplasmic reticulum membrane while HPA describes nuclear staining (UniProt Q6ZMG9 subcellular location; HPA tissue IHC). Use the same controls and scoring rules across compared sections (standard IHC practice).
When is apparent CERS6 positivity credible rather than an artefact?
Give greater weight to reproducible cellular staining with low signal in matched no primary controls and preserved tissue morphology (standard IHC interpretation). Check compartment explicitly: CERS6 is annotated at the endoplasmic reticulum membrane, whereas HPA reports nuclear tissue staining with low RNA staining consistency (UniProt Q6ZMG9 subcellular location; HPA tissue IHC). HPA reports high staining in cerebellar granular layer cells and no detected staining in adipocytes from adipose tissue, but these observations are context rather than universal controls (HPA tissue IHC). Treat staining restricted to section edges, necrotic areas or sites of residual endogenous enzyme activity as suspect (standard chromogenic IHC interpretation). Corroborate an unexpected compartment or cell pattern with an independent validation method before assigning it to CERS6 (standard IHC validation practice).
Boster reagents
Best CERS6 / Ceramide synthase 6 IHC Antibodies
A07033 has IHC-P data from mouse brain tissue and IF data from mouse brain cells (catalog image captions). Its listed reactivity covers human, mouse and rat (catalog: reactivity).
Real IHC data
Anti-LASS6 CERS6 Antibody
Cat # A07033
A07033 lists IHC-P and IF applications, with an IHC image of mouse brain tissue at 2.5 μg/mL (catalog: applications; IHC image caption). Its IF image shows mouse brain cells at 5 μg/mL; the catalog lists human, mouse and rat reactivity (catalog: IF image caption; reactivity).
Which to pick: Choose A07033 for paraffin-section IHC because IHC-P is listed and its own IHC image shows staining in mouse brain tissue; the image caption does not report the fixative (catalog: applications; IHC image caption). For IF, A07033 has a mouse brain-cell image, but ICC validation is unreported (catalog: IF image caption; applications). For work across species, A07033 lists human, mouse and rat reactivity; the supplied IHC and IF images show mouse samples only, and clonality is unreported (catalog: reactivity; image captions; clone field).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q6ZMG9 (CERS6_HUMAN, Ceramide synthase 6).
- Human Protein Atlas. CERS6 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. CERS6 subcellular location (ICC-IF): Variation in transcript expression is not correlated to the cell cycle..
- Human Protein Atlas. CERS6 antibody validation summary (1 antibodies).
- CERS6 promotes esophageal squamous cell carcinoma proliferation by increasing the stability of RPN1. Cell death discovery 2025 — PMC12594893.
- CERS6 required for cell migration and metastasis in lung cancer. Journal of cellular and molecular medicine 2020 — PMC7579715.
- TGF-β receptor I/II trafficking and signaling at primary cilia are inhibited by ceramide to attenuate cell migration and tumor metastasis. Science signaling 2017 — PMC5818989.
- By activating Fas/ceramide synthase 6/p38 kinase in lipid rafts, stichoposide D inhibits growth of leukemia xenografts. Oncotarget 2015 — PMC4695011.
- PubMed PMID:14702039 — UniProt-cited evidence.
- PubMed PMID:15815621 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for CERS6 (Ceramide synthase 6). Expected localisation Nuclear in tissues (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: A07033 at 2.5 μg/mL (the product's own IHC validation image is shown). Controls (HPA): positive Cerebellum (Cells in granular layer); negative Adipose tissue. 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 A07033); 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-CERS6, 2.5 μg/mL (datasheet A07033); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across sections. (standard IHC practice; not target-specific); Caveat: Antibody staining has low concordance with RNA (HPA tissue IHC); Isoform / epitope: 2 isoforms; epitope coverage needs checking (UniProt); Regulation: Staining induction unreported (UniProt). Evidence: UniProt Q6ZMG9, HPA tissue IHC · Approved, HPA cell-line RNA · Variation in transcript expression is not correlated to the cell…, Datasheet A07033, PMC12594893, PMC4695011. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/CERS6/immunohistochemistry-ihc-guide.