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ADGRG6 / Adhesion G protein-coupled receptor G6 · IHC design guide
Design Immunohistochemistry for ADGRG6
Plan ADGRG6 staining in paraffin sections using endothelial cells as a tissue-level reference (HPA tissue IHC). This guide covers the expected membrane location and receptor cleavage (UniProt), plus the catalog antibody’s 1:100–1:300 IHC dilution (datasheet A30803).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
ADGRG6 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 | Cell membrane expected (UniProt); endothelial cells stain (HPA tissue IHC) | |
| Staining pattern | Endothelial cells stain; compartment unresolved (HPA tissue IHC) | |
| Antigen retrieval | Tris-EDTA pH 9.0 HIER, heat-mediated (datasheet A30803) | |
| Positive control | Skeletal muscle+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 | Staining and RNA show low consistency (HPA tissue IHC) | |
| Regulation | Surface detection in activated cells (UniProt) | |
| Isoform / epitope | 4 isoforms; cleavage makes N- vs C-terminal epitope key (UniProt) |
Section 1
Recommended ADGRG6 IHC & IF Protocols
The catalog antibody’s IHC-P protocol (datasheet: A30803) is accompanied by published ADGRG6 IHC protocols from bladder tumors (PMC6372626) and mouse spine sections (PMC8328515).
| Sample | Paraffin-embedded human liver cancer tissue; fixative not specified (datasheet A30803) |
| Fixation | Image fixative and duration unreported (datasheet A30803); 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 9.0 (datasheet A30803); 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-ADGRG6, 1:100 - 1:300 (datasheet A30803) |
| 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 | ADGRG6-positive staining in myocytes of skeletal muscle (HPA tissue IHC: High). HPA tissue profile: Highly expressed in endothelial cells. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated Tris-EDTA at pH 9.0 (datasheet: A30803); the mouse spine protocol also reports Tris-EDTA at pH 9.0 (PMC8328515).
Section 2
What Is the Expected ADGRG6 Staining Pattern?
ADGRG6 is a cell membrane receptor with 7 transmembrane segments (UniProt Q86SQ4 topology). In paraffin section IHC, assess staining in myocytes and smooth muscle cells, where HPA reports high signal, and in endothelial cells, where its tissue profile emphasizes expression (HPA: tissue IHC). HPA rates the IHC evidence Approved but reports low consistency between antibody staining and RNA expression (HPA: reliability description).
| Defined staining at cell borders in skeletal muscle myocytes or smooth muscle cells. | This fits the expected membrane compartment (UniProt Q86SQ4: cell membrane) and HPA's High staining in both cell types (HPA: skeletal muscle myocytes; smooth muscle cells). Assess cell borders against the counterstain and surrounding tissue; a uniformly colored section cannot establish membrane localisation (general IHC practice). |
| Endothelial staining is visible, while adjacent cells stain differently or remain pale. | An endothelial pattern is plausible (HPA: highly expressed in endothelial cells); cerebral cortex endothelial cells are Medium and colon endothelial cells Low in the supplied observations (HPA: tissue IHC). Do not require every vessel to match: UniProt reports ADGRG6 in aortic but not skin microvascular endothelial cells and on activated but not resting umbilical vein cell surfaces (UniProt Q86SQ4: tissue specificity; subcellular location). |
| Strong nuclear staining dominates, without convincing cell border staining. | A nuclear dominant result conflicts with the annotated membrane location (UniProt Q86SQ4: cell membrane; topology). Treat it as suspect rather than as proof of a new ADGRG6 compartment; compare the positive control and a detection control before assigning the signal to the target (general IHC practice). |
| Cells annotated as negative stain as strongly as the expected positive cells. | For example, HPA reports adipocytes and adrenal gland glandular cells as Not detected (HPA: tissue IHC). Signal in those specific cells warrants a specificity check; antibody cross reactivity or endogenous detection activity are possible causes, rather than established explanations for an individual slide (general IHC practice). |
| Diffuse color obscures cell boundaries, or the known positive cell population has no signal. | Diffuse color prevents a defensible localisation call (general IHC practice). An absent result in skeletal muscle myocytes or smooth muscle cells conflicts with HPA's High observations (HPA: tissue IHC), but does not alone prove biological absence or a particular preparation failure. |
💡Expected ADGRG6 appearanceCall a result positive when cell associated, preferably border enriched staining is distinguishable in HPA High myocytes or smooth muscle cells, with plausible endothelial signal where present (UniProt Q86SQ4: membrane; HPA: tissue IHC); isolated nuclear color or uniform haze is suspect (general IHC practice).
| Compartment and epitope position | UniProt places residues 38–862 extracellularly and describes 7 transmembrane segments with cytoplasmic regions (UniProt Q86SQ4: topology). The supplied record gives no antibody epitope, so topology supports a membrane expectation but cannot predict which receptor segment the IHC antibody detects. |
| Proteolytic processing | UniProt describes extracellular N terminal and membrane bound C terminal fragments that predominantly remain associated (UniProt Q86SQ4: processing; subunit, by similarity). Interpret border staining with this architecture in mind; the record does not establish that a specific IHC pattern reflects fragment separation or shedding. |
| Cell type and endothelial state | HPA grades skeletal and smooth muscle cells High, cerebral cortex endothelium Medium, and colon endothelium Low (HPA: tissue IHC). UniProt's endothelial observations differ by vessel source and activation state (UniProt Q86SQ4: tissue specificity; subcellular location); do not treat every unstained vessel as assay failure. |
| Isoforms and antibody evidence | UniProt lists 4 isoforms (UniProt Q86SQ4: isoforms). The supplied HPA antibody record lists HPA017346 as IHC Approved and gives no ICC validation or isoform specific epitope information (HPA: antibody record). Neither isoform recognition nor an IHC Enhanced validation claim follows from these data. |
| Agreement with RNA and tissue controls | HPA calls the IHC reliability Approved while noting low consistency with RNA and staining in unannotated cells or structures (HPA: reliability description). Interpret the pictured cell type directly; an RNA enriched tissue designation for liver or placenta is not a substitute for a cell resolved IHC positive control (HPA: RNA specificity; tissue IHC). |
| IF/ICC Q: what location should be expected? | A: Membrane is the supported location (HPA: subcellular summary; UniProt Q86SQ4: cell membrane). HPA supplies no ICC IF images or ICC antibody validation in this payload (HPA: subcellular record; antibody record), so it does not establish an IF staining pattern. Use the separate IF/ICC guide for that application. |
| Situation | Likely cause | Next action |
|---|---|---|
| No signal in a section expected to contain skeletal muscle myocytes or smooth muscle cells. | HPA reports High staining in those cells (HPA: tissue IHC); absent color may reflect a failed stain or a sampling or cell identification issue (general IHC practice). | Confirm the expected cells are present on the counterstained section, then review the IHC antibody and detection controls and the run's retrieval and detection steps (general IHC practice). No ADGRG6 specific retrieval condition is supplied. |
| Endothelial cells are pale or negative while a muscle control stains. | The endothelial result is context dependent: cortex endothelial cells are Medium and colon endothelial cells Low (HPA: tissue IHC); UniProt also reports vessel source and activation differences (UniProt Q86SQ4). | Identify the vessel and cell population before calling failure; compare with the run's positive control and HPA's matched tissue observation, where available (HPA: tissue IHC; general IHC practice). |
| Nuclei are strongly colored but cell borders are not. | Predominant nuclear staining is inconsistent with the membrane annotation (UniProt Q86SQ4: subcellular location). The cause cannot be assigned from compartment alone. | Check a detection control and the positive tissue for the same nuclear pattern, then reassess antibody specificity and interpretation (general IHC practice). Do not score nuclear color alone as ADGRG6 positive. |
| Adipocytes or adrenal gland glandular cells show convincing color. | Those cell populations are Not detected in the supplied HPA observations (HPA: tissue IHC). Cross reactivity or endogenous detection activity is possible (general IHC practice). | Compare a reagent omission control and the expected positive cells on the same run; review endogenous activity blocking as appropriate for chromogenic detection (general IHC practice). |
| Color spreads broadly across tissue and masks cell boundaries. | Nonspecific background or detection activity can obscure compartment assignment (general IHC practice); HPA's membrane summary cannot validate diffuse color (HPA: subcellular summary). | Inspect reagent omission and detection controls, blocking, washing, and development time; score only signal that can be assigned to cells and location (general IHC practice). |
| Two specimens show different distributions or staining strengths. | HPA reports uneven cell type levels and low antibody staining to RNA consistency (HPA: tissue IHC; reliability description). UniProt also lists 4 isoforms, without an epitope map for this antibody (UniProt Q86SQ4: isoforms; HPA: antibody record). | Compare the same cell types, compartments, and run controls before calling a biological difference; avoid attributing discordance to an isoform or fixation effect without separate evidence (general IHC practice). |
Experiment evidence
IHC & IF images showing ADGRG6 staining
A30803 · IHCImmunohistochemical analysis of paraffin-embedded human liver cancer. 1, Antibody was diluted at 1:200(4° overnight). 2, Tris-EDTA,pH9.0 was
A32344-1 · IHCImmunohistochemical analysis of paraffin-embedded human liver cancer. 1, Antibody was diluted at 1:200 (4° overnight). 2, Tris-EDTA, pH9.0 w
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000112414-ADGRG6/tissue/cerebral+cortex#imid_4497052
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000112414-ADGRG6/tissue/colon#imid_4496970
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000112414-ADGRG6/tissue/liver#imid_4496963
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000112414-ADGRG6/tissue/kidney#imid_4496980
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for ADGRG6 IHC & IF
🧪Run skeletal muscle first and look for staining in myocytes (HPA: High in myocytes). Use adipose tissue as a negative and expect adipocytes to remain at background (HPA: Not detected in adipocytes); on the skeletal-muscle slide, treat unstained nonmyocyte cells as internal background only after confirming their identity, since the supplied HPA row does not classify them as negative (HPA: High in myocytes).
Positive control tissue: Skeletal muscle (Myocytes, 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 ADGRG6; derive a cell-line control from the positive tissue's cell type (Myocytes) and confirm it by RNA or western blot first.
Technical controls: Include a no-primary, secondary-only control and an isotype control matched to the primary antibody’s host species and clonality; use ADGRG6 knockout tissue, if available, as a biological specificity control (standard IHC practice). For chromogenic staining of skeletal muscle, quench endogenous peroxidase and compare signal with the no-primary slide (standard chromogenic IHC practice).
⚠️Feasibility: No target-specific fixation window or fixation effect is reported in the supplied evidence, and the selected paraffin-section caption does not state a fixative (catalog antibody A30803 tissue-IHC caption). That caption uses Tris-EDTA at pH 9.0 for retrieval and a 1:200 primary dilution overnight at 4°C; it does not establish that retrieval is required (catalog antibody A30803 tissue-IHC caption). Neither frozen sections nor IF are shown to be easier in the supplied evidence; in skeletal muscle, endogenous peroxidase can complicate chromogenic interpretation (standard IHC practice).
HPA tissue IHC evidence for ADGRG6
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Low consistency between antibody staining and RNA expression data. Antibody staining in cells/structures not annotated, view images.). 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 |
|---|---|---|---|---|
| Skeletal muscle | Myocytes | High | Protein (IHC) | HPA → |
| Smooth muscle | Smooth muscle cells | High | Protein (IHC) | HPA → |
| Breast | Myoepithelial cells | Medium | Protein (IHC) | HPA → |
| Cerebral cortex | Endothelial cells | Medium | Protein (IHC) | HPA → |
| Heart muscle | Cardiomyocytes | 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 → |
| Appendix | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | Not detected | Protein (IHC) | HPA → |
| Caudate | Glial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced ADGRG6 IHC Tips
Troubleshoot ADGRG6 staining in paraffin section IHC using the catalog antibody’s tissue image, receptor topology, and cell specific reference staining.
Which retrieval conditions should I try first for weak ADGRG6 staining?
Start with heat mediated retrieval in Tris-EDTA at pH 9.0 for paraffin sections (datasheet A30803). The catalog image used this retrieval with antibody at 1:200 overnight at 4°C in paraffin embedded human liver cancer (caption A30803). If staining is weak, compare a shorter and longer heating interval on adjacent sections while holding antibody concentration and detection constant (standard IHC practice). Check tissue integrity and background before increasing retrieval intensity, since damaged sections can make a stronger signal difficult to interpret (standard IHC practice). Score vascular and other identifiable cell compartments separately rather than judging total brown area alone (HPA: highly expressed in endothelial cells).
Can fixation explain weak or uneven ADGRG6 staining?
The paraffin section image does not report its fixative, and ADGRG6 specific sensitivity to fixation is unknown (caption A30803). Record the fixative, fixation duration, section age, and processing history for each sample before comparing staining across batches (standard IHC practice). Keep those variables consistent when testing the stated Tris-EDTA pH 9.0 retrieval and 1:200 primary dilution (datasheet A30803; caption A30803). If staining differs between batches, repeat adjacent sections in one staining run with the same detection and development times (standard IHC practice). Do not attribute a fixation effect to receptor cleavage, glycosylation, or tissue expression without a direct comparison (UniProt Q86SQ4 processing and glycosylation).
Where should convincing ADGRG6 staining appear in tissue sections?
Prioritize staining along cell boundaries because ADGRG6 is a cell membrane receptor (UniProt Q86SQ4 subcellular location; HPA subcellular: membrane). Examine endothelial profiles carefully, while identifying the stained cell type from morphology and an appropriate companion marker (HPA: highly expressed in endothelial cells; standard IHC practice). Strong myocyte and smooth muscle cell staining has also been reported in tissue IHC, so vascular staining is not the only plausible pattern (HPA: high in skeletal muscle myocytes and smooth muscle cells). Compare the distribution with a negative reagent control and avoid calling diffuse nuclear color receptor localization without independent support (standard IHC practice; UniProt Q86SQ4 topology).
How could epitope placement change the ADGRG6 IHC pattern?
Confirm the antibody’s mapped epitope before assigning a stained structure to a receptor fragment, since the supplied caption does not specify an epitope (caption A30803). ADGRG6 has 4 isoforms and can yield an extracellular N terminal fragment spanning residues 38–840 and a membrane bound C terminal fragment spanning 841–1221 (UniProt Q86SQ4 isoforms and processing). Its large extracellular region spans residues 38–862, while the distal cytoplasmic region spans 1114–1221 (UniProt Q86SQ4 topology). An antibody recognizing one side of the cleavage site may report a different distribution from one recognizing the other, so compare interpretations with epitope documentation and appropriate controls (UniProt Q86SQ4 processing; standard IHC practice).
How should I assess ADGRG6 in multiplex immunofluorescence?
Treat IF as a separate assay: the supplied tissue image establishes paraffin section IHC conditions, and HPA lists no cell line ICC/IF images here (caption A30803; HPA subcellular record). Pair ADGRG6 with an endothelial marker when testing vascular localization, then confirm that the signals share the same cells and membrane contours (HPA: highly expressed in endothelial cells; UniProt Q86SQ4 subcellular location). Choose a fluorophore channel after checking tissue autofluorescence with an unstained section and single color controls (standard IF practice). For a verified extracellular epitope, first assess staining without permeabilisation; for a verified cytoplasmic epitope, test permeabilisation while monitoring membrane integrity (UniProt Q86SQ4 topology; standard IF practice).
What should I check when ADGRG6 staining looks diffuse or excessively dark?
First compare a no primary control with the stained section to assess detection background, and inspect tissue edges and damaged areas separately (standard IHC practice). For chromogenic detection, verify the peroxidase block and shorten chromogen development if widespread brown signal persists (standard IHC practice). The catalog image used a 1:200 primary overnight at 4°C and a 1:200 secondary for 45 minutes at room temperature; these are reference conditions for that image, not proof of specificity (caption A30803). Reassess any diffuse cytoplasmic or nuclear signal against the expected membrane localization and identifiable cell outlines (UniProt Q86SQ4 subcellular location; standard IHC practice).
How can I quantify ADGRG6 IHC across sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the compartment and cell population before scoring, because ADGRG6 is membrane associated and endothelial staining can contribute to a section’s total signal (UniProt Q86SQ4 subcellular location; HPA: highly expressed in endothelial cells). For a defined cell population, report the percentage of positive cells and a prespecified intensity score, or calculate an H-score from those categories (standard IHC practice). For vessels, report positive endothelial profiles per mm² of viable tissue and normalize to vessel density or total endothelial profiles measured with a companion marker (standard IHC practice). Use the same threshold, section area rules, retrieval, and chromogen development across cases, and exclude folds and necrotic regions before analysis (standard IHC practice).
How do I distinguish true ADGRG6 positivity from IHC artefact?
Look for reproducible cell boundary staining in morphologically intact cells, consistent with ADGRG6 membrane localization (UniProt Q86SQ4 subcellular location; standard IHC practice). An endothelial pattern is plausible, but assign cell identity from morphology or a companion marker rather than brown color alone (HPA: highly expressed in endothelial cells; standard IHC practice). Treat edge restricted staining, necrotic debris, and signal also present in a no primary control as possible artefacts; check peroxidase blocking when enzyme related background is suspected (standard IHC practice). Interpret a positive result cautiously because HPA rates its tissue staining approved while reporting low consistency with RNA expression and staining in unannotated cells or structures (HPA tissue IHC reliability description).
Boster reagents
Best ADGRG6 / Adhesion G protein-coupled receptor G6 IHC Antibodies
Both anti-ADGRG6 antibodies have IHC images from paraffin-embedded human liver cancer (catalog image captions). Both list Human and Mouse reactivity; A30803 also lists Rat (catalog reactivity).
Real IHC data
Anti-GPR126 ADGRG6 Antibody
Cat # A30803
Real IHC data
Anti-GPR126 ADGRG6 Antibody
Cat # A32344-1
A30803 will render with its paraffin-embedded human liver cancer IHC image; it lists IHC, IF and ICC applications and Human, Mouse and Rat reactivity (catalog image caption; applications/reactivity). A32344-1 will render with its own paraffin-embedded human liver cancer IHC image; it lists IHC and IF applications and Human and Mouse reactivity (catalog image caption; applications/reactivity).
Which to pick: For tissue IHC, either A30803 or A32344-1 has its own paraffin-section image using 1:200 primary antibody and Tris-EDTA retrieval at pH 9.0; neither caption reports the fixative (each SKU’s IHC image caption). For IF/ICC, choose A30803 when ICC is needed because it lists both applications, while A32344-1 lists IF only; both are rabbit polyclonals (catalog applications; host; dilution_raw). For Rat samples, choose A30803, which lists Rat reactivity; A32344-1 lists Human and Mouse (catalog reactivity).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q86SQ4 (AGRG6_HUMAN, Adhesion G protein-coupled receptor G6).
- Human Protein Atlas. ADGRG6 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. ADGRG6 subcellular location (ICC-IF): Membrane.
- Human Protein Atlas. ADGRG6 antibody validation summary (1 antibodies).
- Characterization of ADGRG6 as a potential molecular oncotarget of pancreatic cancer. Cell death & disease 2026 — PMC13316050.
- Whole-genome sequencing identifies ADGRG6 enhancer mutations and FRS2 duplications as angiogenesis-related drivers in bladder cancer. Nature communications 2019 — PMC6372626.
- An adhesion G protein-coupled receptor is required in cartilaginous and dense connective tissues to maintain spine alignment. eLife 2021 — PMC8328515.
- The G protein-coupled receptor ADGRG6 maintains mouse growth plate homeostasis through IHH signaling. Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research 2024 — PMC11523133.
- PubMed PMID:15225624 — UniProt-cited evidence.
- PubMed PMID:15189448 — UniProt-cited evidence.
- PubMed PMID:17974005 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for ADGRG6 (Adhesion G protein-coupled receptor G6). Expected localisation Cell membrane expected (UniProt); endothelial cells stain (HPA tissue IHC). Antigen retrieval: Tris-EDTA pH 9.0 HIER, heat-mediated (datasheet A30803). Validated IHC antibody: A30803 at 1:100 - 1:300 (the product's own IHC validation image is shown). Controls (HPA): positive Skeletal muscle (Myocytes); negative Adipose tissue. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Paraffin-embedded human liver cancer tissue; fixative not specified (datasheet A30803); 2. Fixation: Image fixative and duration unreported (datasheet A30803); 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 retrieval: Tris-EDTA pH 9.0 (datasheet A30803); 20 min, 95–100 °C (standard); 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-ADGRG6, 1:100 - 1:300 (datasheet A30803); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across sections. (standard IHC practice; not target-specific); Caveat: Staining and RNA show low consistency (HPA tissue IHC); Isoform / epitope: 4 isoforms; cleavage makes N- vs C-terminal epitope key (UniProt); Regulation: Surface detection in activated cells (UniProt). Evidence: UniProt Q86SQ4, HPA tissue IHC · Approved, HPA ICC-IF · Membrane, Datasheet A30803, PMC6372626, PMC8328515. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/ADGRG6/immunohistochemistry-ihc-guide.