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ADCY6 / Adenylate cyclase type 6 · IHC design guide
Design Immunohistochemistry for ADCY6
Plan ADCY6 paraffin IHC around cytoplasmic staining in most tissues (HPA tissue IHC). Use colon glandular cells as a positive reference and oral mucosal squamous cells as a negative reference, while checking the catalog antibody’s ADCY5/6 specificity (HPA tissue IHC; datasheet: A05655-1).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
ADCY6 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 | 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 | Adrenal gland+4 more · see all | |
| Negative control | Adipose tissue+2 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep formalin fixation consistent across sections (standard IHC practice; not target-specific) | |
| Caveat | Catalog antibody is labeled ADCY5/6; verify specificity (datasheet: A05655-1) | |
| Regulation | Expression regulation unreported (UniProt) | |
| Isoform / epitope | 2 isoforms; cytoplasmic versus extracellular epitope unknown (UniProt) |
Section 1
Recommended ADCY6 IHC & IF Protocols
The catalog antibody’s IHC-P protocol is accompanied by one published ADCY6 IHC protocol (PMC9377375).
| Sample | Paraffin-embedded human brain tissue; fixative not specified (datasheet A05655-1) |
| Fixation | Image fixative and duration unreported (datasheet A05655-1); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat-induced epitope retrieval in citrate buffer, pH 6.0, 20 min at 95–98 °C (standard rule: cytoplasmic / membrane antigen) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% normal serum of the secondary host, 30 min, room temperature (standard) |
| Primary antibody | Rabbit anti-ADCY6, 1:100-1:300 (datasheet A05655-1) |
| 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 | ADCY6-positive staining in glandular cells of adrenal gland (HPA tissue IHC: Medium). HPA tissue profile: Cytoplasmic expression in most tissues. No signal in the no-primary control. |
💡Decision noteStart with citrate pH 6.0 retrieval at 95–98 °C for 20 min (page retrieval rule); the published protocol uses 100 °C for 16 min (PMC9377375).
Section 2
What Is the Expected ADCY6 Staining Pattern?
ADCY6 is a multi-pass membrane protein at the cell membrane, cilia and stereocilia (UniProt O43306: localisation and 12 transmembrane segments). In tissue IHC, expect predominantly cytoplasmic staining in glandular and other positive cell populations (HPA tissue IHC: cytoplasmic expression in most tissues). Interpret that pattern cautiously: HPA rates the IHC antibody Approved but reports low consistency between antibody staining and RNA expression (HPA tissue IHC: reliability description).
| Cytoplasmic staining in glandular cells of adrenal gland, appendix, colon or duodenum. | This fits the reported tissue pattern; each listed glandular population has Medium staining (HPA tissue IHC). Judge signal within the named cells, since HPA's tissue observation does not establish that every neighboring cell should stain (HPA tissue IHC: cell-specific entries). |
| Cytoplasmic staining in bronchial respiratory epithelial cells, marrow hematopoietic cells or cerebral cortex endothelial cells. | These are additional Medium-staining populations (HPA tissue IHC). Their presence can support an IHC result, while differences among tissues need cautious interpretation because antibody staining and RNA expression show low consistency (HPA tissue IHC: reliability description). |
| Strong nuclear-only staining, with little staining in the expected cellular compartment. | Treat this as a localisation mismatch requiring controls, rather than a convincing ADCY6 pattern: tissue IHC is described as cytoplasmic (HPA tissue IHC), and UniProt places ADCY6 at membranes and cell projections (UniProt O43306: subcellular location). |
| Prominent staining in adipocytes, oral squamous epithelium or ovarian stromal cells. | Those populations are reported Not detected (HPA tissue IHC). Check whether the signal is nonspecific antibody binding or detection background before assigning it to ADCY6; a single discordant field cannot by itself identify the cause (HPA tissue IHC: negative entries). |
| Diffuse staining across tissue and blank areas, or no staining in a listed Medium-positive population. | The first pattern suggests background in the staining run; the second calls for a positive-control and detection check (standard IHC practice). Neither establishes biological absence, especially with the reported low IHC–RNA consistency (HPA tissue IHC: reliability description). |
💡Expected ADCY6 appearanceCall positive when cytoplasmic signal is discernible in a listed Medium-positive cell population, such as colon glandular cells (HPA tissue IHC); widespread blank-area or nuclear-only colour is a suspect positive pattern (standard IHC interpretation; HPA tissue IHC: cytoplasmic profile).
| Compartment assignment | ADCY6 has 12 transmembrane segments and annotated membrane, cilium and stereocilium locations (UniProt O43306); HPA nevertheless describes tissue IHC as cytoplasmic (HPA tissue IHC). Record what the section shows without requiring a crisp membrane rim. |
| Choice of comparison tissue and cells | Adrenal, appendix, colon and duodenum glandular cells are Medium, whereas adipocytes, oral squamous cells and ovarian stromal cells are Not detected (HPA tissue IHC). Compare named cell populations, not whole-tissue averages. |
| Strength of IHC evidence | The listed antibody, CAB018365, is IHC Approved; HPA also reports low consistency with RNA expression (HPA antibodies; HPA tissue IHC). An unexpected positive needs corroboration before a biological conclusion. |
| IF/ICC Q: should an IF image match the tissue IHC pattern? | A: HPA reports approved Golgi localisation in ICC-IF images, while its tissue IHC summary is cytoplasmic; CAB018365 has ICC status Uncertain (HPA subcellular; HPA tissue IHC; HPA antibodies). Interpret the two applications separately. |
| Situation | Likely cause | Next action |
|---|---|---|
| No stain in colon or appendix glandular cells. | A listed Medium-positive population is missing, but the image alone cannot distinguish a failed run from variation in the sampled section (HPA tissue IHC). | Confirm that the relevant glands are present, review the positive control, then check retrieval and detection steps as general IHC troubleshooting (standard IHC practice). |
| Nuclear-only signal dominates. | The compartment conflicts with HPA's cytoplasmic tissue profile and UniProt's membrane and cell-projection annotations (HPA tissue IHC; UniProt O43306). | Inspect secondary-only or other suitable detection controls and compare a listed positive tissue before scoring the nuclear signal (standard IHC practice; HPA tissue IHC). |
| Adipocytes or oral squamous cells stain strongly. | Those cells are listed Not detected; nonspecific binding or detection background is possible, though the image cannot identify which (HPA tissue IHC; standard IHC interpretation). | Compare the suspect cells with a listed Medium-positive population and review antibody-omission and detection controls (HPA tissue IHC; standard IHC practice). |
| Brown colour is diffuse or extends into blank areas. | A noncellular or poorly bounded reaction is consistent with staining background (standard chromogenic IHC interpretation). | Review blocking, wash and detection controls, including an antibody-omission control; score only cell-associated signal that can be assigned to a tissue population (standard IHC practice). |
| Signal appears only at cell borders, without the reported cytoplasmic pattern. | A border signal could reflect ADCY6's annotated membrane location, but it differs from HPA's cytoplasmic tissue IHC summary (UniProt O43306; HPA tissue IHC). | Document the compartment and compare named positive and Not detected cell populations; do not treat topology alone as proof that the border stain is specific (HPA tissue IHC; UniProt O43306). |
| An IF Golgi image seems inconsistent with the IHC section. | HPA assigns Golgi localisation in ICC-IF but describes cytoplasmic staining in tissue IHC; the listed antibody's ICC status is Uncertain (HPA subcellular; HPA tissue IHC; HPA antibodies). | Evaluate each application against its own HPA observation and validation status, then base this section's tissue call on IHC controls and cell-specific staining (HPA tissue IHC; HPA antibodies). |
Experiment evidence
IHC & IF images showing ADCY6 staining
A05655-1 · IHCImmunohistochemistry analysis of paraffin-embedded human brain tissue, using ADCY5/6 Antibody. The picture on the right is blocked with the
A05655-1 · IFImmunofluorescence analysis of HeLa cells, using ADCY5/6 Antibody. The picture on the right is blocked with the synthesized peptide.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000174233-ADCY6/tissue/cerebral+cortex#imid_4587960
HPA · CerebellumHuman Protein Atlas IHC, Cerebellum. Source: https://www.proteinatlas.org/ENSG00000174233-ADCY6/tissue/cerebellum#imid_4587971
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000174233-ADCY6/tissue/colon#imid_4587902
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000174233-ADCY6/tissue/liver#imid_4587895
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody CAB018365; green = ADCY6, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody CAB018365; green = ADCY6, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for ADCY6 IHC & IF
🧪Run bronchus first: respiratory epithelial cells should show ADCY6 staining (HPA: Medium in bronchus respiratory epithelial cells). Use oral mucosa squamous epithelial cells as the negative tissue (HPA: Not detected); on the bronchus slide, any independently validated negative cells should show no specific chromogen deposition above background (standard IHC practice).
Positive control tissue: Adrenal gland (Glandular cells, HPA Medium)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show ADCY6 in A-431, U-251MG, U2OS, with annotated localisation: Golgi apparatus (approved) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control; a host-species- and isotype-matched control IgG appropriate to the antibody's clonality; and an ADCY6 knockout sample or cognate-peptide block (standard IHC practice). Quench endogenous peroxidase and check inflammatory cells in bronchus for residual chromogen background (standard IHC practice).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected paraffin-section caption does not state a fixative (caption: A05655-1). Retrieval dependence is unreported; compare antigen-retrieval conditions on matched sections before scoring (standard IHC practice). Frozen sections and tissue IF cannot be ranked as easier from the supplied evidence; HPA reports ICC-IF images with approved Golgi localization, while bronchial inflammatory cells warrant a check for endogenous-peroxidase background in chromogenic IHC (HPA subcellular; standard IHC practice).
HPA tissue IHC evidence for ADCY6
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 |
|---|---|---|---|---|
| Adrenal gland | Glandular cells | Medium | Protein (IHC) | HPA → |
| Appendix | Glandular cells | Medium | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | Medium | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | Medium | Protein (IHC) | HPA → |
| Cerebral cortex | Endothelial cells | Medium | Protein (IHC) | HPA → |
Section 3
Advanced ADCY6 IHC Tips
Troubleshoot ADCY6 chromogenic IHC in paraffin sections by checking retrieval, compartment, cell type, antibody specificity and scoring controls.
How should I retrieve ADCY6 antigen in paraffin sections?
Start with citrate buffer at pH 6.0 for heat-induced retrieval at 95–98 °C for 20 min (page retrieval rule). Keep section thickness, heating vessel and cooling conditions consistent across the run, then compare staining with a section processed without primary antibody (standard IHC practice). If staining remains weak, test a second retrieval condition on adjacent sections while retaining citrate pH 6.0 as the reference condition (standard IHC practice). Judge improvement by cell-specific staining and background together; ADCY6 has 12 transmembrane segments, so widespread nuclear signal warrants scrutiny rather than stronger retrieval alone (UniProt O43306 topology).
Could fixation explain weak or uneven ADCY6 staining?
Target-specific fixation sensitivity is unknown because the supplied paraffin-section caption does not state a fixative or fixation time (A05655-1 tissue-IHC caption). Record each specimen’s fixative and processing history, and compare adjacent sections only when these conditions are matched (standard IHC practice). If staining is weak, compare controlled fixation conditions using the same citrate pH 6.0, 95–98 °C, 20 min retrieval baseline (page retrieval rule; standard IHC practice). Evaluate tissue preservation and staining distribution together; neither the HPA staining pattern nor ADCY6 topology establishes a fixation effect for this antibody (HPA tissue IHC; UniProt O43306 topology).
What localisation should count as plausible ADCY6 staining?
Assess chromogenic signal against cell boundaries and cytoplasm: ADCY6 is annotated at the cell membrane, cilium and stereocilium, while HPA tissue IHC describes cytoplasmic expression (UniProt O43306 subcellular; HPA tissue IHC). HPA ICC/IF additionally reports an approved Golgi location, which can help frame a compact perinuclear pattern but does not validate that pattern in paraffin IHC (HPA subcellular). Compare cells within the same tissue compartment and inspect whether staining tracks intact structures rather than folds or torn edges (standard IHC practice). Predominantly nuclear deposits should trigger review of detection background and antibody specificity before being scored as ADCY6 (UniProt O43306 subcellular; standard IHC practice).
How do isoforms and epitope position affect this IHC result?
ADCY6 has 2 annotated isoforms, but the supplied evidence does not map this antibody’s epitope or establish isoform selectivity (UniProt O43306 isoforms; A05655-1 tissue-IHC caption). Check whether the antibody documentation identifies a peptide sequence and whether that sequence is shared by both isoforms before assigning an isoform-specific score (standard IHC practice). ADCY6 includes cytoplasmic regions at residues 1–151, 306–673 and 914–1168, plus an extracellular region at 760–819; access depends on the actual epitope (UniProt O43306 topology). The caption describes an ADCY5/6 antibody and peptide-blocked image, so blocked staining alone cannot establish ADCY6-specific recognition (A05655-1 tissue-IHC caption).
How can I cross-check an IHC pattern using multiplex IF?
On a separate IF/ICC workflow, pair ADCY6 with a validated endothelial-cell marker when examining cerebral cortex, where HPA reports medium endothelial staining (HPA tissue IHC; standard IF practice). Choose spectrally separated fluorophores after checking tissue autofluorescence and include single-stain controls to identify bleed-through (standard IF practice). For a cytoplasmic epitope, optimise mild permeabilisation; an extracellular epitope at residues 760–819 may be assessed before permeabilisation if the antibody recognises it (UniProt O43306 topology; standard IF practice). Because the supplied antibody epitope is unmapped, compare permeabilised and nonpermeabilised preparations before interpreting discordant IF and chromogenic IHC patterns (A05655-1 tissue-IHC caption; standard IF practice).
How do I reduce diffuse or granular DAB background?
Run no-primary and reagent controls, then inspect whether deposits follow tissue edges, damaged areas or endogenous pigment rather than intact cells (standard IHC practice). Use an appropriate peroxidase block before DAB development and shorten development if controls accumulate colour; these are general chromogenic IHC steps, not ADCY6-specific validation (standard IHC practice). Titrate primary antibody and blocking conditions on matched sections while keeping citrate pH 6.0 retrieval fixed at first (page retrieval rule; standard IHC practice). Compare the resulting pattern with reported cytoplasmic tissue expression and membrane or projection annotations, while remembering that HPA reports low staining-to-RNA consistency (HPA tissue IHC; UniProt O43306 subcellular).
How should I score ADCY6 staining across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, then record the percentage of positive cells and staining intensity in those cells (standard IHC practice). An H-score can combine intensity with percentage; use identical intensity thresholds and DAB development conditions across sections (standard IHC practice). For spatial comparisons, report positive-cell density per mm² of viable tissue and normalise to the area or number of evaluable cells in the specified compartment (standard IHC practice). Keep glandular and endothelial populations separate where relevant, because HPA assigns ADCY6 staining to different cell types across tissues and reports low staining-to-RNA consistency (HPA tissue IHC).
When is an ADCY6-positive IHC result convincing?
A convincing result has reproducible staining in intact cells, a plausible cytoplasmic or membrane-associated pattern, and low signal in the no-primary control (HPA tissue IHC; UniProt O43306 subcellular; standard IHC practice). Check cell identity: HPA reports medium staining in cerebral-cortex endothelial cells and no detection in adipocytes, but its tissue IHC has low consistency with RNA expression (HPA tissue IHC). Exclude edge effects, necrotic areas and endogenous peroxidase deposits before calling a cell positive (standard IHC practice). The peptide-blocked brain image supports peptide-sensitive staining for the labeled ADCY5/6 antibody; it does not by itself distinguish ADCY6 from ADCY5 (A05655-1 tissue-IHC caption).
Boster reagents
Best ADCY6 / Adenylate cyclase type 6 IHC Antibodies
A05655-1 has IHC data from paraffin-embedded human brain tissue and IF data from HeLa cells (catalog image captions); its listed reactivity is human, mouse and rat (catalog applications/reactivity).
Real IHC data
Anti-A Cyclase V/VI ADCY6 Antibody
Cat # A05655-1
A05655-1 will render with an IHC image of paraffin-embedded human brain tissue, including a peptide-blocked comparison (catalog IHC caption). It is also listed for IF, with an image of HeLa cells and a peptide-blocked comparison (catalog applications; catalog IF caption).
Which to pick: For tissue IHC, choose A05655-1: its own image shows paraffin-embedded human brain tissue, but the fixative is unreported (catalog IHC caption). For cell IF/ICC, A05655-1 has IF data in HeLa cells; ICC is not separately listed (catalog IF caption; catalog applications). For cross-species work, A05655-1 is a rabbit polyclonal listed as reactive with human, mouse and rat, while the supplied IHC image documents human tissue only (catalog dilution_raw/reactivity; catalog IHC caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry O43306 (ADCY6_HUMAN, Adenylate cyclase type 6).
- Human Protein Atlas. ADCY6 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. ADCY6 subcellular location (ICC-IF): Localized to the Golgi apparatus..
- Human Protein Atlas. ADCY6 antibody validation summary (1 antibodies).
- ADCY6 is a potential prognostic biomarker and suppresses OTSCC progression via Hippo signaling pathway. The Kaohsiung journal of medical sciences 2023 — PMC11895994.
- MiR-27a-3p binds to TET1 mediated DNA demethylation of ADCY6 regulates breast cancer progression via epithelial-mesenchymal transition. Frontiers in oncology 2022 — PMC9377375.
- Decreased microRNA-182-5p helps alendronate promote osteoblast proliferation and differentiation in osteoporosis via the Rap1/MAPK pathway. Bioscience reports 2018 — PMC6435538.
- PubMed PMID:10978539 — UniProt-cited evidence.
- PubMed PMID:9455477 — UniProt-cited evidence.
- PubMed PMID:12168954 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for ADCY6 (Adenylate cyclase type 6). Expected localisation 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: A05655-1 at 1:100-1:300 (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Glandular cells); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human brain tissue; fixative not specified (datasheet A05655-1); 2. Fixation: Image fixative and duration unreported (datasheet A05655-1); verify before use.; 3. Sectioning: 4–5 µm sections on charged slides (standard); 4. Deparaffinisation: Xylene, graded ethanol series to water (standard); 5. Antigen retrieval: Heat-induced epitope retrieval in citrate buffer, pH 6.0 , 20 min at 95–98 °C…; 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% normal serum of the secondary host, 30 min , room temperature (standard); 8. Primary antibody: Rabbit anti-ADCY6, 1:100-1:300 (datasheet A05655-1); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep formalin fixation consistent across sections (standard IHC practice; not target-specific); Caveat: Catalog antibody is labeled ADCY5/6; verify specificity (datasheet: A05655-1); Isoform / epitope: 2 isoforms; cytoplasmic versus extracellular epitope unknown (UniProt); Regulation: Expression regulation unreported (UniProt). Evidence: UniProt O43306, HPA tissue IHC · Approved, HPA ICC-IF · Golgi apparatus., Datasheet A05655-1, PMC9377375. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/ADCY6/immunohistochemistry-ihc-guide.