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CMTM6 / CKLF-like MARVEL transmembrane domain-containing protein 6 · IHC design guide
Design Immunohistochemistry for CMTM6
Plan chromogenic CMTM6 IHC in paraffin sections using the predominantly cytoplasmic tissue pattern (HPA tissue IHC). Start with the catalog antibody at 1:100–1:300 (datasheet), and compare high-staining endometrial glandular cells with undetected staining in skeletal myocytes (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
CMTM6 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 | Predominantly cytoplasmic tissue staining (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic staining in most 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 | Hippocampus+1 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep paraffin-section fixation consistent across samples (standard IHC practice; not target-specific) | |
| Caveat | Staining has medium consistency with RNA expression (HPA tissue IHC) | |
| Regulation | Expression regulation unspecified (UniProt) | |
| Isoform / epitope | No isoforms reported; epitope side matters across 4 transmembrane spans (UniProt) |
Section 1
Recommended CMTM6 IHC & IF Protocols
The catalog antibody’s IHC-P protocol is accompanied by published CMTM6 protocols for gastric cancer, hepatocellular carcinoma, and renal cell carcinoma (PMC7842018; PMC8353897; PMC13039633).
| Sample | Paraffin-embedded human oophoroma tissue; fixative not specified (datasheet A14107) |
| Fixation | Image fixative and duration unreported (datasheet A14107); 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-CMTM6, 1:100-1:300 (datasheet A14107) |
| 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 | CMTM6-positive staining in purkinje cells of cerebellum (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in most tissues. No signal in the no-primary control. |
💡Decision noteStart with citrate pH 6.0 at 95–98 °C for 20 min (page retrieval rule). The gastric protocol used EDTA pH 9 (PMC7842018).
Section 2
What Is the Expected CMTM6 Staining Pattern?
CMTM6 is a four-pass membrane protein found at the cell membrane and in early and recycling endosomes (UniProt Q9NX76 topology and subcellular location). In tissue IHC, expect chiefly cytoplasmic staining, with signal in glandular cells, urothelial cells and Purkinje cells among the HPA high-staining examples (HPA tissue IHC). HPA rates its tissue staining reliability Enhanced, while reporting medium consistency between antibody staining and RNA expression (HPA tissue IHC).
| Cytoplasmic staining in endometrial, fallopian tube or rectal glandular cells. | This matches HPA's chiefly cytoplasmic tissue profile and High staining in these cell types (HPA tissue IHC). A cytoplasmic appearance can be compatible with endosomal membrane localization; IHC alone does not identify the stained organelle (UniProt Q9NX76 subcellular location; general IHC practice). |
| Membrane-associated signal, possibly alongside cytoplasmic signal, in cells with expected staining. | A membrane component is compatible with CMTM6 at the cell membrane and on early and recycling endosome membranes (UniProt Q9NX76 subcellular location). Judge it alongside HPA's observed cytoplasmic tissue pattern; a membrane-only result is not the sole required appearance (HPA tissue IHC). |
| Predominantly nuclear staining, with little cytoplasmic or membrane signal. | A nuclear-dominant pattern does not match the reported cell membrane, endosomal or tissue IHC locations (UniProt Q9NX76 subcellular location; HPA tissue IHC). Treat it as suspect and assess background and antibody specificity before scoring cells as CMTM6 positive (general IHC practice). |
| Strong staining in hippocampal glial cells or skeletal myocytes. | HPA reports CMTM6 as Not detected in those respective cells, so strong signal is discordant with these reference observations (HPA tissue IHC). Cross-reactivity or endogenous detection activity are possibilities to investigate, although one discordant slide does not establish either cause (general IHC practice). |
| No signal in a tissue containing a documented high-staining cell population. | Absence in, for example, endometrial glandular cells or urinary bladder urothelial cells conflicts with HPA's High observations (HPA tissue IHC). First check that the relevant cells are present and assess the staining run; HPA's Enhanced reliability does not guarantee every specimen will stain (HPA tissue IHC; general IHC practice). |
💡Expected CMTM6 appearanceCall a result consistent with CMTM6 when the appropriate cells show discernible, chiefly cytoplasmic staining, with a possible membrane component and stronger signal in HPA High populations; isolated nuclear signal or strong staining in HPA Not detected cell types is suspect (HPA tissue IHC; UniProt Q9NX76 subcellular location).
| Cell population and reference intensity | HPA reports High staining in Purkinje cells, several glandular populations and urinary bladder urothelial cells, but Not detected staining in hippocampal glial cells and skeletal myocytes (HPA tissue IHC). Compare like cell types when choosing and reading controls; whole-tissue labels can obscure which cells matter (general IHC practice). |
| Compartment and topology | CMTM6 has four transmembrane segments and reported cell membrane and endosomal membrane locations (UniProt Q9NX76 topology and subcellular location). HPA describes tissue staining as cytoplasmic, so interpret visible cytoplasmic signal in that context without claiming that chromogenic IHC resolves a specific endosome (HPA tissue IHC; general IHC practice). |
| Evidence and antibody validation | HPA calls tissue IHC reliability Enhanced and notes medium agreement between staining and RNA data; HPA026980 has Enhanced IHC validation (HPA tissue IHC; HPA antibodies). These support a reference pattern, while unexpected compartments or cell types still warrant controls and review (general IHC practice). |
| Detection background | Endogenous detection activity and nonspecific reagent staining can produce chromogen unrelated to primary-antibody binding (general IHC practice). A no-primary control helps assess detection-system background; it cannot by itself establish that remaining signal is specific to CMTM6 (general IHC practice). |
| IF/ICC question | Q: What localization should an IF/ICC image be compared with? A: HPA reports mainly vesicular CMTM6 signal and an Approved ICC result for HPA063491; UniProt also reports cell membrane and recycling endosome localization (HPA subcellular; HPA antibodies; UniProt Q9NX76 subcellular location). Use the separate IF/ICC guide for its workflow. |
| Situation | Likely cause | Next action |
|---|---|---|
| Expected high-staining cells are blank. | The target cells may be absent from the section, or a staining-run step may have failed; the image alone cannot distinguish these explanations (general IHC practice). | Confirm the relevant cell population and run a documented positive tissue alongside the sample; review the IHC-validated antibody's specified workflow and the run's detection controls (HPA tissue IHC; general IHC practice). |
| Most tissue compartments show a diffuse brown haze. | Broad signal without cell or compartment definition is consistent with nonspecific staining or detection background (general IHC practice). | Compare with a no-primary control, inspect wash and blocking steps, and reassess the signal only after background is separated from the cellular pattern (general IHC practice). |
| Strong signal is concentrated in nuclei. | This location conflicts with reported membrane and endosomal localization and the HPA cytoplasmic tissue profile (UniProt Q9NX76 subcellular location; HPA tissue IHC). | Check whether the nuclear signal persists in controls, then evaluate primary-antibody specificity before including it in a positive score (general IHC practice). |
| Hippocampal glia or skeletal myocytes stain strongly. | Those cell types are listed as Not detected by HPA; cross-reactivity or endogenous detection activity may account for discordant signal (HPA tissue IHC; general IHC practice). | Verify the cell identity, compare a no-primary control and review staining in a documented High cell population within the same run (HPA tissue IHC; general IHC practice). |
| Only an apparent membrane rim is visible. | Cell membrane localization is supported, but a rim alone does not reproduce HPA's chiefly cytoplasmic tissue profile (UniProt Q9NX76 subcellular location; HPA tissue IHC). | Inspect the expected cell populations at suitable magnification and compare the complete cellular pattern with a positive tissue control before assigning a compartment-based score (HPA tissue IHC; general IHC practice). |
| Signal varies sharply between otherwise comparable sections. | Target-specific fixation effects are not established by the supplied assay evidence. Verify with a matched IHC source before attributing a result to fixation. | Compare matching cell populations and concurrent controls, then review the recorded IHC workflow for run differences without attributing the variation to a target-specific fixation effect (general IHC practice). |
Experiment evidence
IHC & IF images showing CMTM6 staining
A14107 · IHCImmunohistochemical analysis of paraffin-embedded human-oophoroma, antibody was diluted at 1:100
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000091317-CMTM6/tissue/cerebral+cortex#imid_6838768
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000091317-CMTM6/tissue/colon#imid_6838694
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000091317-CMTM6/tissue/liver#imid_6838687
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000091317-CMTM6/tissue/kidney#imid_6838680
HPA · ICC-IF RT-4Human Protein Atlas immunofluorescence, RT-4, antibody HPA063491; green = CMTM6, blue = nucleus, red = microtubules.
HPA · ICC-IF RT-4Human Protein Atlas immunofluorescence, RT-4, antibody HPA063491; green = CMTM6, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for CMTM6 IHC & IF
🧪Run urinary bladder first: urothelial cells should stain (HPA: High in urothelial cells). Use skeletal muscle as the negative tissue, where myocytes are not detected (HPA: Not detected in myocytes); on the bladder slide, cells independently verified as CMTM6-negative should show only background staining (standard IHC practice).
Positive control tissue: Cerebellum (Purkinje cells, HPA High)
Negative control tissue: Hippocampus (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show CMTM6 in RT-4, U-251MG, U2OS, with annotated localisation: Vesicles (approved) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control and a control antibody matched to the primary antibody’s host species and clonality: an isotype match for a monoclonal antibody or nonimmune IgG for a polyclonal antibody (standard IHC practice). Confirm specificity with a CMTM6-knockout biological control processed in parallel, and block endogenous peroxidase on bladder sections before chromogenic detection (standard IHC practice).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported in the supplied evidence; the A14107 paraffin-section caption gives a 1:100 dilution but does not report the fixative (selected-SKU tissue-IHC caption: fixative not stated). Retrieval dependence is unreported, so assess antigen retrieval empirically for the IHC-validated antibody (standard IHC practice); the evidence does not establish whether frozen sections or IF are easier. In bladder, distinguish urothelial staining from signal in any leukocytes present, which may express CMTM6 (HPA: High in urothelial cells; UniProt Q9NX76: expressed in leukocytes).
HPA tissue IHC evidence for CMTM6
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Enhanced — Medium consistency between antibody staining and RNA expression data.). Rows are taken directly from the HPA tissue chart — click any row's HPA link to view the source.
Positive expression · recommended positive controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Cerebellum | Purkinje cells | High | Protein (IHC) | HPA → |
| Endometrium | Glandular cells | High | Protein (IHC) | HPA → |
| Fallopian tube | Glandular cells | High | Protein (IHC) | HPA → |
| Rectum | Glandular cells | High | Protein (IHC) | HPA → |
| Urinary bladder | Urothelial cells | High | Protein (IHC) | HPA → |
Section 3
Advanced CMTM6 IHC Tips
Troubleshoot CMTM6 staining in paraffin sections by checking retrieval, tissue context, subcellular pattern and controls before scoring chromogenic signal.
How should I retrieve CMTM6 antigen when paraffin sections stain weakly?
Start with citrate buffer at pH 6.0, heated to 95–98 °C for 20 min (page retrieval rule). Let sections cool in buffer, then compare staining with a matched section processed in the same run (standard IHC practice). CMTM6 has 4 transmembrane segments and short extracellular loops, so epitope accessibility may depend on the antibody’s binding site (UniProt Q9NX76 topology). If staining remains weak, vary heating time on replicate sections while keeping the buffer and detection conditions constant (standard IHC practice). Check tissue morphology and background before accepting stronger signal as improved retrieval (standard IHC practice).
How can I troubleshoot fixation when CMTM6 staining varies between sections?
The selected image shows staining in paraffin-embedded human oophoroma, but its caption does not report a fixative (A14107 tissue-IHC caption). Target-specific sensitivity to fixation is therefore unknown; neither tissue staining patterns nor protein topology establish a suitable fixation duration (A14107 tissue-IHC caption; UniProt Q9NX76 topology). Record the fixative and processing history for each specimen, and compare sections with matched processing when possible (standard IHC practice). Use citrate at pH 6.0 for 20 min at 95–98 °C across that comparison (page retrieval rule). If signal differs, assess morphology and controls before attributing the difference to fixation (standard IHC practice).
Should CMTM6 appear on cell surfaces or in the cytoplasm?
Evaluate both cell borders and intracellular puncta: CMTM6 is reported at the cell membrane and early and recycling endosome membranes (UniProt Q9NX76 subcellular location). It colocalizes with PD-L1 at the plasma membrane and recycling endosomes, while the subcellular atlas reports mainly vesicular localization (UniProt Q9NX76 subcellular location; HPA subcellular). Tissue IHC is described as cytoplasmic in most tissues, so a cytoplasmic chromogenic pattern alone is plausible but does not identify its organelle (HPA tissue IHC). Compare staining at 1:100 only when using the selected antibody under comparable conditions (A14107 tissue-IHC caption). Review cell boundaries and puncta at high magnification, alongside a negative control (standard IHC practice).
How does the CMTM6 epitope affect interpretation of a weak stain?
Ask which region the antibody recognizes before treating weak staining as low protein abundance (standard IHC practice). The supplied record lists 0 isoforms and a 183-residue chain, so it provides no isoform-specific explanation for staining differences (UniProt Q9NX76 isoforms and processing). Its 4 membrane-spanning segments separate cytoplasmic regions from short extracellular loops, making the epitope’s location relevant to accessibility (UniProt Q9NX76 topology). The record also lists phosphorylation at residues 8 and 171, but supplies no evidence that either modification changes this antibody’s staining (UniProt Q9NX76 modified residues). Compare retrieval conditions and tissue controls before assigning a molecular cause (standard IHC practice).
How should I check a CMTM6 pattern by multiplex IF?
Use the separate IF/ICC workflow to pair CMTM6 with a marker for the expected cell type, then inspect signal within those cells (standard IF practice). For example, glandular cells are reported as high in endometrium, whereas skeletal myocytes are not detected in the tissue atlas (HPA tissue IHC). Choose fluorophores in channels with low measured tissue autofluorescence, and include single-label controls to assess bleed-through (standard IF practice). Set permeabilisation according to the antibody epitope: cytoplasmic CMTM6 regions require intracellular access, while extracellular loops face outward at the plasma membrane (UniProt Q9NX76 topology; standard IF practice). Check for membrane and vesicular signal without assuming complete overlap with PD-L1 (UniProt Q9NX76 subcellular location; standard IF practice).
What should I change when CMTM6 chromogenic staining is diffuse?
First compare the stained section with a no-primary control and inspect whether color follows tissue edges, damaged areas or endogenous pigment (standard IHC practice). Block endogenous peroxidase before horseradish peroxidase detection, and keep development time consistent between sections (standard chromogenic IHC practice). If background persists, titrate the primary around the captioned 1:100 dilution for SKU A14107 and adjust blocking or washing one variable at a time (A14107 tissue-IHC caption; standard IHC practice). Genuine signal may be cytoplasmic in tissue and vesicular at subcellular resolution, so diffuse color is not automatically false (HPA tissue IHC; HPA subcellular). Confirm that any improvement preserves staining in expected cells (HPA tissue IHC; standard IHC practice).
How should I score CMTM6 across heterogeneous tissue sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartments before scoring, because tissue IHC is commonly cytoplasmic and CMTM6 is also reported at membranes and vesicles (HPA tissue IHC; UniProt Q9NX76 subcellular location). For an H-score, multiply each intensity grade 0–3 by its percentage of cells and sum to a 0–300 score (standard IHC scoring practice). Alternatively, report percentage of positive cells or positive-cell density per mm², with the threshold fixed across the batch (standard IHC scoring practice). Normalize each measure to eligible, intact cells or analyzed tissue area, as appropriate (standard IHC scoring practice). Record epithelial and infiltrating-cell results separately where both populations are present (standard IHC scoring practice).
When is apparent CMTM6 positivity likely to be an artefact?
Favor a pattern in intact cells that matches the expected population and includes plausible cytoplasmic, membrane or vesicular signal (HPA tissue IHC; UniProt Q9NX76 subcellular location; HPA subcellular). High staining is reported in endometrial glandular cells, whereas skeletal myocytes are reported as not detected, providing context for tissue-specific comparisons (HPA tissue IHC). Treat nuclear-only color, staining confined to section edges or necrosis, and signal reproduced in a no-primary control as reasons to investigate artefact (standard IHC practice). Check peroxidase blocking when chromogen appears without primary antibody (standard chromogenic IHC practice). CMTM6 associates with PD-L1, but CMTM6 positivity alone does not measure PD-L1 expression or inhibitory signaling (UniProt Q9NX76 function; standard IHC interpretation).
Boster reagents
Best CMTM6 / CKLF-like MARVEL transmembrane domain-containing protein 6 IHC Antibodies
A14107 is a human- and mouse-reactive anti-CMTM6 antibody listed for IHC and IF (catalog: applications and reactivity). Its IHC image shows a paraffin-embedded human oophoroma section (A14107 IHC image caption).
Real IHC data
Anti-CMTM6 Antibody
Cat # A14107
A14107 will render with IHC data from a paraffin-embedded human oophoroma section at 1:100 (A14107 IHC image caption). The catalog lists A14107 for human and mouse IHC and IF, but supplies no IF image (catalog: applications, reactivity, and image captions).
Which to pick: Choose A14107 for paraffin-section IHC because its own image caption documents that preparation; the fixative is unreported (A14107 IHC image caption). For IF/ICC planning, A14107 lists IF at 1:50, but ICC validation and an IF image are unreported (catalog: applications, IF dilution, and image captions). A14107 is the listed cross-species option because the catalog reports human and mouse reactivity; it is a rabbit polyclonal antibody (catalog: reactivity, host, and dilution_raw).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q9NX76 (CKLF6_HUMAN, CKLF-like MARVEL transmembrane domain-containing protein 6).
- Human Protein Atlas. CMTM6 tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. CMTM6 subcellular location (ICC-IF): Mainly localized to vesicles..
- Human Protein Atlas. CMTM6 antibody validation summary (2 antibodies).
- Co-expression of CMTM6 and PD-L1: a novel prognostic indicator of gastric cancer. Cancer cell international 2021 — PMC7842018.
- High membrane expression of CMTM6 in hepatocellular carcinoma is associated with tumor recurrence. Cancer science 2021 — PMC8353897.
- Molecular and immunological characteristics of patients with CMTM6 low expression colorectal cancer. Medicine 2023 — PMC10727655.
- Association of CMTM6 expression with clinicopathological characteristics and prognostic implications in renal cell carcinoma. Therapeutic advances in medical oncology 2026 — PMC13039633.
- PubMed PMID:12782130 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for CMTM6 (CKLF-like MARVEL transmembrane domain-containing protein 6). Expected localisation Predominantly cytoplasmic tissue staining (HPA tissue IHC). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: A14107 at 1:100-1:300 (the product's own IHC validation image is shown). Controls (HPA): positive Cerebellum (Purkinje cells); negative Hippocampus. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Paraffin-embedded human oophoroma tissue; fixative not specified (datasheet A14107); 2. Fixation: Image fixative and duration unreported (datasheet A14107); 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-CMTM6, 1:100-1:300 (datasheet A14107); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep paraffin-section fixation consistent across samples (standard IHC practice; not target-specific); Caveat: Staining has medium consistency with RNA expression (HPA tissue IHC); Isoform / epitope: No isoforms reported; epitope side matters across 4 transmembrane spans (UniProt); Regulation: Expression regulation unspecified (UniProt). Evidence: UniProt Q9NX76, HPA tissue IHC · Enhanced, HPA ICC-IF · Mainly localized to vesicles., Datasheet A14107, PMC7842018, PMC8353897. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/CMTM6/immunohistochemistry-ihc-guide.