CERKL / Ceramide kinase-like protein · IHC design guide

Design Immunohistochemistry for CERKL

Plan CERKL paraffin IHC using cytoplasmic tissue staining and high signal in lung alveolar cells as reference points (HPA tissue IHC). This guide covers the catalog antibody’s 1:100–1:300 IHC dilution (datasheet A09801-1) and the uncertain agreement between staining and RNA expression (HPA tissue IHC).

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

Immunohistochemistry protocol sheet for CERKL (IHC for CERKL): expected localisation Cytoplasm (HPA tissue IHC); nucleolar enrichment is a molecular expectation (UniProt), antibody A09801-1, validated IHC image, and IHC protocol steps — Printable **CERKL** IHC protocol sheet — expected localisation Cytoplasm (HPA tissue IHC); nucleolar enrichment is a molecular expectation (UniProt), antibody A09801-1, controls and protocol steps. Open the full CERKL IHC guide →

Navigate to: Protocols Staining pattern Controls Antibodies

CERKL 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	Cytoplasm (HPA tissue IHC); nucleolar enrichment is a molecular expectation (UniProt)
	Staining pattern	Cytoplasmic in most tissues; high in lung alveolar cells (HPA tissue IHC)
	Antigen retrieval	Tris-EDTA pH 8.0 HIER, heat-mediated (datasheet A09801-1)
	Positive control ⓘ	Adrenal gland+4 more · see all
	Negative control ⓘ	Heart muscle+4 more · see all

Important caveats

Reasons your staining may differ from the expected pattern.

	Fixation	Keep fixation conditions consistent across sections. (standard IHC practice; not target-specific)
	Caveat	Staining–RNA agreement is uncertain (HPA tissue IHC)
	Regulation	Isoform expression varies by tissue (UniProt)
	Isoform / epitope	8 isoforms; epitope coverage is unknown (UniProt)

Section 1

Recommended CERKL IHC & IF Protocols

The catalog antibody’s IHC-P protocol is accompanied by two published CERKL IHC methods (PMC8172666; PMC12645229).

Recommended immunohistochemistry (IHC-P) protocol parameters
Sample	Paraffin-embedded Human breast cancer tissue; fixative not specified (datasheet A09801-1)
Fixation	Image fixative and duration unreported (datasheet A09801-1); verify before use.
Sectioning	4–5 µm sections on charged slides (standard)
Deparaffinisation	Xylene, graded ethanol series to water (standard)
Antigen retrieval	Heat retrieval: Tris-EDTA pH 8.0 (datasheet A09801-1); 20 min, 95–100 °C (standard)
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-CERKL, 1:100-1:300 (datasheet A09801-1)
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	CERKL-positive staining in glandular cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in most tissues. No signal in the no-primary control.

💡Decision noteStart with heat-mediated Tris-EDTA retrieval at pH 8.0 for the catalog antibody (datasheet A09801-1).

Section 2

What Is the Expected CERKL Staining Pattern?

CERKL is mainly cytoplasmic in tissue IHC, with high staining reported in glandular cells, lung alveolar cells, and other listed cell populations (HPA: tissue IHC). Nuclear staining can also fit its annotated localisation (UniProt Q49MI3; HPA: approved ICC-IF nucleoplasm). CERKL has no transmembrane segment (UniProt Q49MI3 topology). Treat tissue intensity as provisional: HPA rates its IHC profile Uncertain because antibody staining and RNA expression have low consistency (HPA: tissue IHC).

What am I looking at on my slide?
Cytoplasmic staining in pancreatic exocrine glandular cells, lung alveolar cells, or colonic glandular cells.	This matches cell populations scored High by tissue IHC (HPA: pancreas, lung, colon). Compare the cellular distribution with a negative control; high HPA staining is a reference pattern, not proof that an individual antibody signal is specific (HPA: IHC reliability Uncertain; standard IHC practice).
Nuclear staining accompanies cytoplasmic staining in otherwise plausible cells.	Nuclear signal can be biologically plausible: nucleoplasm and cytosol are approved ICC-IF locations (HPA: subcellular), and UniProt annotates nuclear and nucleolar localisation (UniProt Q49MI3). Tissue IHC is described chiefly as cytoplasmic, so assess nuclear detail and control staining before assigning it to CERKL (HPA: tissue IHC).
Signal is confined to cell membranes or extracellular material, with no convincing intracellular staining.	That pattern does not fit the reported cytoplasmic tissue profile or the annotated intracellular locations (HPA: tissue IHC; UniProt Q49MI3). Check the negative control and tissue morphology for nonspecific deposits or detection artefacts before interpreting the signal as CERKL (standard IHC practice).
Strong staining appears in cardiomyocytes or skeletal myocytes while expected comparison cells remain weak.	HPA reports CERKL as Not detected in those cell populations (HPA: heart muscle, skeletal muscle). This mismatch raises concern about cross-reactivity or endogenous detection activity; HPA's Uncertain IHC reliability means the discrepancy warrants controls rather than a definitive specificity judgment (HPA: tissue IHC; standard IHC practice).
Broad, even chromogen obscures cell borders and the negative control also stains.	A shared background pattern prevents reliable compartment or cell-type scoring (standard IHC practice). Review blocking, washing, and endogenous enzyme controls for the chosen detection system, then compare the corrected slide with HPA's predominantly cytoplasmic reference pattern (standard IHC practice; HPA: tissue IHC).

💡Expected CERKL appearanceA positive tissue IHC result shows distinct cytoplasmic signal in an HPA High cell population, such as pancreatic exocrine glandular cells; isolated membranous or extracellular deposits are suspect, and HPA rates the tissue profile Uncertain (HPA: pancreas and tissue IHC; UniProt Q49MI3 topology).

How each factor affects the staining
Tissue and cell selection	HPA reports High staining in pancreatic exocrine glandular cells and lung alveolar cells, but Not detected in cardiomyocytes and skeletal myocytes (HPA: tissue IHC). Compare named cell populations within intact tissue rather than treating every cell in a positive tissue as positive (standard IHC practice).
Compartment and isoform context	UniProt lists cytoplasm, nucleus, nucleolus, Golgi, and endoplasmic reticulum, with nucleolar enrichment except for isoform 5 (UniProt Q49MI3). HPA approves nucleoplasm and cytosol in ICC-IF, while tissue IHC is predominantly cytoplasmic (HPA: subcellular and tissue IHC).
Antibody evidence	HPA rates the overall tissue IHC profile Uncertain and lists HPA035444 as IHC Uncertain; neither status establishes a definitive positive or negative result for another antibody (HPA: tissue IHC; HPA: antibodies). Use the tested antibody's own controls when judging a section (standard IHC practice).
Chromogenic detection background	Endogenous enzyme activity can contribute color when an enzyme-based IHC detection system is used (standard IHC practice). A detection-only control helps separate that activity from primary-antibody staining; HPA's tissue pattern does not identify the source of background in a particular slide (standard IHC practice; HPA: tissue IHC).

Why is my staining missing, weak or wrong?
Situation	Likely cause	Next action
No stain in pancreatic exocrine cells or lung alveolar cells.	Either the assay failed or this specimen differs from HPA's High reference pattern; HPA rates its IHC profile Uncertain (HPA: pancreas, lung, tissue IHC).	Confirm morphology and run a suitable positive control, then check primary-antibody dilution, retrieval, and detection steps using the antibody's validated IHC instructions (standard IHC practice).
Cardiomyocytes or skeletal myocytes stain strongly.	These cells are Not detected in HPA tissue IHC, so cross-reactivity or chromogenic background is possible (HPA: heart muscle, skeletal muscle; standard IHC practice).	Compare primary-omission and detection controls with the stained section; judge the suspect cells separately from adjacent tissue (standard IHC practice).
Only sharp membrane or extracellular staining is visible.	The distribution conflicts with CERKL's intracellular annotations and predominantly cytoplasmic tissue profile (UniProt Q49MI3; HPA: tissue IHC).	Inspect morphology and control slides for edge deposits or nonspecific signal; do not score that pattern alone as CERKL positivity (standard IHC practice).
Nuclear signal is present but cytoplasmic signal is weak.	Nuclear localisation is plausible, although HPA describes tissue IHC chiefly as cytoplasmic (UniProt Q49MI3; HPA: subcellular and tissue IHC).	Compare nuclear staining with the negative control and tissue cell types before scoring; report the observed compartment rather than forcing a cytoplasmic-only call (standard IHC practice).
Diffuse color makes intensity or localisation impossible to score.	Background may arise from incomplete blocking, inadequate washing, or endogenous detection activity in a chromogenic assay (standard IHC practice).	Review the detection-only control, blocking, and wash conditions; repeat scoring only where cell boundaries and compartments are discernible (standard IHC practice).
Two sections give different proportions of stained cells.	Cell composition may differ between sections, and HPA reports low consistency between antibody staining and RNA expression (standard IHC practice; HPA: tissue IHC).	Compare the same named cell population in both sections, document its compartment and intensity, and avoid treating HPA's High designation as a universal threshold (standard IHC practice; HPA: tissue IHC).

Source: UniProt Q49MI3 Human Protein Atlas tissue Human Protein Atlas subcellular

Experiment evidence

IHC & IF images showing CERKL staining

Immunohistochemical analysis of paraffin-embedded Human breast cancer. Antibody was diluted at 1:100 (4° overnight). High-pressure and temperature Tris-EDTA, pH8.0 was used for antigen retrieval. Negetive contrl (right) obtaned from antibody was pre-absorbed by immunogen peptide.

A09801-1 · IHCImmunohistochemical analysis of paraffin-embedded Human breast cancer. Antibody was diluted at 1:100 (4° overnight). High-pressure and tempe

HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000188452-CERKL/tissue/cerebral+cortex#imid_8622808

HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000188452-CERKL/tissue/colon#imid_8617622

HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000188452-CERKL/tissue/liver#imid_8617607

HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000188452-CERKL/tissue/kidney#imid_8617624

HPA ICC-IF BJ [Human fibroblast] — CERKL (HPA035443)

HPA · ICC-IF BJ [Human fibroblast]Human Protein Atlas immunofluorescence, BJ [Human fibroblast], antibody HPA035443; green = CERKL, blue = nucleus, red = microtubules.

← swipe / scroll for more images →

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

Sample controls for CERKL IHC & IF

🧪Run colon first: glandular cells should stain strongly (HPA: High in colon glandular cells). Use skeletal muscle as the negative tissue, where myocytes are listed as not detected (HPA: Not detected in skeletal muscle myocytes); on the colon slide, non-glandular cells should show only background staining, but their CERKL-negative status is not established by the supplied HPA row.

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

Negative control tissue: Heart muscle (HPA Not detected)

ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show CERKL in BJ [Human fibroblast], Rh30, U2OS, with annotated localisation: Nucleoplasm (approved), Cytosol (approved) (HPA subcellular).

Technical controls: Include a no-primary, secondary-only control and an isotype control matched to the primary antibody’s host species and clonality (standard IHC practice). Use a CERKL knockout sample or immunogen-peptide absorption as a biological specificity control (selected-SKU tissue-IHC caption: peptide-absorbed negative control), and block endogenous peroxidase in colon sections before chromogenic detection (standard IHC practice).

⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected-SKU paraffin-section caption does not state the fixative (selected-SKU tissue-IHC caption). The caption reports high-pressure, high-temperature Tris-EDTA retrieval at pH 8.0, but does not establish that retrieval is required (selected-SKU tissue-IHC caption). The supplied evidence does not establish whether frozen sections or IF are easier; for IF/ICC, nucleoplasm and cytosol are approved locations, and colon autofluorescence should be checked with an unstained section (HPA subcellular; standard IF practice).

HPA tissue IHC evidence for CERKL

Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Uncertain — 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
Adrenal gland	Glandular cells	High	Protein (IHC)	HPA →
Colon	Glandular cells	High	Protein (IHC)	HPA →
Duodenum	Glandular cells	High	Protein (IHC)	HPA →
Gallbladder	Glandular cells	High	Protein (IHC)	HPA →
Lung	Alveolar cells	High	Protein (IHC)	HPA →

Undetected expression · recommended negative controls

Tissue	Cell type	Level	Evidence	Source
Heart muscle	Cardiomyocytes	Not detected	Protein (IHC)	HPA →
Ovary	Ovarian stroma cells	Not detected	Protein (IHC)	HPA →
Skeletal muscle	Myocytes	Not detected	Protein (IHC)	HPA →
Smooth muscle	Smooth muscle cells	Not detected	Protein (IHC)	HPA →
Soft tissue	Fibroblasts	Not detected	Protein (IHC)	HPA →

Section 3

Advanced CERKL IHC Tips

Troubleshoot CERKL staining in paraffin sections by checking retrieval, controls, compartment, and cell type before comparing chromogenic IHC scores.

Which retrieval conditions should I start with for CERKL IHC?

Start with heat-mediated Tris-EDTA retrieval at pH 8.0 for paraffin sections (datasheet A09801-1). The selected CERKL image used high-pressure, high-temperature Tris-EDTA at pH 8.0, followed by antibody at 1:100 overnight at 4 °C (caption A09801-1). Run a retrieval-treated section beside an otherwise matched control so a weak signal can be separated from staining caused by the detection system (standard IHC practice). If staining remains weak, optimise heating time on adjacent sections before trying another retrieval buffer, while checking tissue preservation and control staining (standard IHC practice).

How should I troubleshoot fixation-related loss of CERKL staining?

The selected image identifies paraffin-embedded tissue but does not state its fixative, so CERKL-specific fixation sensitivity is unknown (caption A09801-1). Record the fixative, duration, and processing history for every section being compared, then keep those conditions matched across cases (standard IHC practice). If staining varies between blocks, assess section morphology and a control stain before attributing the difference to CERKL abundance (standard IHC practice). Optimise retrieval on adjacent sections from the affected block while holding the catalog antibody incubation at the documented 1:100, 4 °C overnight condition during that comparison (caption A09801-1; standard IHC practice).

Should CERKL staining be cytoplasmic, nuclear, or nucleolar?

Evaluate cytoplasmic and nuclear staining separately: CERKL is annotated in the cytoplasm and nucleoli, with possible movement between nucleus and cytoplasm (UniProt Q49MI3). It is also annotated at the Golgi apparatus, trans-Golgi network, and endoplasmic reticulum, while approved cell-image locations are nucleoplasm and cytosol (UniProt Q49MI3; HPA subcellular). HPA describes predominantly cytoplasmic tissue staining, but its tissue IHC reliability is uncertain because staining and RNA data have low consistency (HPA tissue IHC). Compare compartment patterns in matched sections and examine whether apparent nuclear signal follows intact nuclei rather than folds, pigment, or precipitate (standard IHC practice).

Could CERKL splice isoforms explain different compartment patterns?

CERKL has 8 listed isoforms, and isoform 5 lacks the nucleolar enrichment reported for CERKL generally (UniProt Q49MI3). Before interpreting a missing nucleolar signal, check which sequence the catalog antibody recognises and whether its epitope is shared by the isoforms relevant to the specimen (UniProt Q49MI3; standard IHC practice). The record places the DAGKc domain at residues 164–339, but supplies no antibody epitope, so domain location alone cannot establish isoform coverage (UniProt Q49MI3). Score nuclear, nucleolar, and cytoplasmic staining separately when comparing specimens, and avoid assigning an isoform from chromogenic localisation alone (standard IHC practice).

How can I check a CERKL IHC pattern with multiplex IF?

Use IF on matched material to examine whether CERKL overlaps a marker identifying the cell population scored by IHC; HPA reports high staining in pancreatic exocrine glandular cells and lung alveolar cells (HPA tissue IHC; standard IF practice). Choose fluorophores after inspecting tissue autofluorescence in an unstained control, and reserve a spectrally distinct channel for the cell-type marker (standard IF practice). CERKL has no transmembrane segment and has cytoplasmic and nuclear annotations, so permeabilise sufficiently to access intracellular epitopes while preserving compartment boundaries (UniProt Q49MI3; standard IF practice). Keep this IF comparison exploratory because the supplied catalog evidence documents paraffin-section IHC conditions, not an IF protocol (caption A09801-1).

What should I check when CERKL DAB staining is diffuse?

First compare the stained section with a no-primary control and inspect whether DAB appears in the same compartments without antibody (standard IHC practice). Block endogenous peroxidase before chromogenic detection, then review wash stringency and detection-reagent background on matched sections (standard IHC practice). The selected image used antibody at 1:100 overnight at 4 °C and included a control pre-absorbed with immunogen peptide, which provides a useful specificity comparison for that image (caption A09801-1). Treat diffuse staining cautiously because HPA describes widespread cytoplasmic tissue expression yet rates its CERKL tissue IHC reliability uncertain (HPA tissue IHC).

How should I quantify CERKL staining across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION

Define the cell population and compartment before scoring, because CERKL can appear in cytoplasmic and nuclear locations (UniProt Q49MI3). For a consistent cell population, record the percentage of positive cells and intensity, then calculate an H-score using percentages at intensity levels 0–3 (standard IHC practice). Alternatively, report positive-cell density per mm² of viable tissue when cell counts vary substantially between fields (standard IHC practice). Normalise comparisons to the same eligible cell type, viable tissue area, staining batch, and exposure or scan settings; report cytoplasmic and nuclear results separately (standard IHC practice).

How do I distinguish credible CERKL staining from artefact?

A credible pattern should occur in intact cells and fit a plausible CERKL compartment: cytoplasm, nucleus, or nucleolus, with isoform 5 as a possible exception to nucleolar enrichment (UniProt Q49MI3). Compare cell identity with the expected tissue pattern, while recognising that HPA tissue IHC has uncertain reliability and low staining–RNA consistency (HPA tissue IHC). Discount signals concentrated at section edges, folds, necrotic regions, or sites that also stain in the no-primary control (standard IHC practice). Check endogenous peroxidase blocking and compare the documented peptide-absorption control when available before calling unexpected DAB deposits CERKL-positive (standard IHC practice; caption A09801-1).

Boster reagents

Best CERKL / Ceramide kinase-like protein IHC Antibodies

One anti-CERKL antibody has a real IHC image from human breast cancer paraffin sections; IF is listed, but no IF image is supplied (catalog applications; IHC image caption).

Real IHC data Immunohistochemical analysis of paraffin-embedded Human breast cancer. Antibody was diluted at 1:100 (4° overnight). High-pressure and temperature Tris-EDTA, pH8.0 was used for antigen retrieval. Negetive contrl (right) obtaned from antibody was pre-absorbed by immunogen peptide.

Anti-Ceramide kinase-like protein CERKL Antibody

Cat # A09801-1

View product →

A09801-1 has an IHC image from human breast cancer paraffin sections (IHC image caption). A09801-1 lists human reactivity and IF among its applications, with no IF image supplied (catalog reactivity and applications; IF image list).

Which to pick: Choose A09801-1 for tissue IHC: its image shows human breast cancer paraffin sections, while the fixative is unreported (IHC image caption). For IF/ICC, A09801-1 lists IF but has no IF image or ICC listing; its host is rabbit and clonality is unreported (catalog applications, IF image list, host and clone). No cross-species option is documented: A09801-1 lists human reactivity only (catalog reactivity).

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

References

UniProt Consortium. UniProt entry Q49MI3 (CERKL_HUMAN, Ceramide kinase-like protein).
Human Protein Atlas. CERKL tissue IHC expression (reliability: Uncertain).
Human Protein Atlas. CERKL subcellular location (ICC-IF): Localized to the nucleoplasm and cytosol..
Human Protein Atlas. CERKL antibody validation summary (3 antibodies).
CERKL is upregulated in cutaneous squamous cell carcinoma and maintains cellular sphingolipids and resistance to oxidative stress. The British journal of dermatology 2021 — PMC8172666.
Spatiotemporal expression pattern of ceramide kinase-like in the mouse retina. Molecular vision 2010 — PMC3000240.
CERKL Reduced PI3P/Autophagy to Promote Pancreatic Cancer. Cancer medicine 2025 — PMC12645229.
CERKL, a Retinal Dystrophy Gene, Regulates Mitochondrial Transport and Dynamics in Hippocampal Neurons. International journal of molecular sciences 2022 — PMC9570143.
PubMed PMID:14681825 — UniProt-cited evidence.
PubMed PMID:15708351 — UniProt-cited evidence.
PubMed PMID:14702039 — UniProt-cited evidence.