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AHCY / Adenosylhomocysteinase · IHC design guide
Design Immunohistochemistry for AHCY
Plan paraffin-section AHCY IHC around nuclear and cytoplasmic staining (HPA tissue IHC). Use kidney tubule cells as a high-staining reference and adipocytes as an undetected reference (HPA tissue IHC), and titrate the catalog antibody within 2–5 μg/ml (datasheet A04236-1).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
AHCY Immunohistochemistry Experimental Design Guide
Expected localisation, validated protocols, controls and antibodies — the at-a-glance facts below, then the full design guide.
Must know before staining
| Expected localisation | Nuclear and cytoplasmic staining (HPA tissue IHC) | |
| Staining pattern | Widespread nuclear and cytoplasmic cellular staining (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A04236-1) | |
| Positive control | Epididymis+4 more · see all | |
| Negative control | Adipose tissue |
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 medium consistency (HPA tissue IHC) | |
| Regulation | Specific regulators unreported (UniProt) | |
| Isoform / epitope | 2 isoforms; epitope coverage is unknown (UniProt) |
Section 1
Recommended AHCY IHC & IF Protocols
The catalog antibody’s IHC-P protocol (datasheet A04236-1) is accompanied by one published AHCY staining protocol for paraffin sections (PMC9345734).
| Sample | Paraffin-embedded human breast cancer tissue; fixative not specified (datasheet A04236-1) |
| Fixation | Image fixative and duration unreported (datasheet A04236-1); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat retrieval: EDTA pH 8.0 (datasheet A04236-1); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A04236-1) |
| Primary antibody | Rabbit anti-AHCY, 2-5 μg/ml (datasheet A04236-1) |
| Primary incubation | Overnight at 4 °C (datasheet A04236-1) |
| Detection | HRP-conjugated secondary, DAB chromogen (datasheet A04236-1) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | AHCY-positive staining in glandular cells of epididymis (HPA tissue IHC: High). HPA tissue profile: Ubiquitous nuclear and cytoplasmic expression. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA pH 8.0 retrieval for the catalog antibody (datasheet A04236-1); the published protocol does not specify retrieval (PMC9345734).
Section 2
What Is the Expected AHCY Staining Pattern?
AHCY should show nuclear and cytoplasmic staining across many tissue cell types, with particularly strong staining in selected glandular, tubular, seminiferous and urothelial cells (HPA tissue IHC). HPA rates the tissue pattern Approved, with medium consistency between staining and RNA data (HPA tissue IHC). AHCY has no transmembrane segment, so a membrane-restricted pattern needs scrutiny (UniProt P23526 topology).
| Nuclear and cytoplasmic staining in kidney tubule cells, with stronger signal than nearby weakly stained cells. | This fits the reported high tubule-cell signal and broad nuclear–cytoplasmic distribution (HPA tissue IHC). Judge intensity within the same section; ubiquitous expression does not mean every cell stains equally (HPA tissue IHC). |
| Signal confined to cell membranes or extracellular spaces, without convincing intracellular staining. | That distribution conflicts with HPA tissue IHC and the absence of a transmembrane segment (UniProt P23526 topology). Review morphology and controls before calling it AHCY; UniProt also lists nucleus and endoplasmic reticulum as locations (UniProt P23526). |
| Strong adipocyte staining, or dominant signal in a cell population expected to stain weakly. | Adipocytes were not detected in the HPA tissue panel, while glial, alveolar and muscle cells had low signal (HPA tissue IHC). Unexpected staining may reflect cross-reactivity or endogenous detection activity; compare with appropriate controls (general IHC practice). |
| Uniform colour over cells and empty spaces, obscuring cell boundaries. | Diffuse background prevents a reliable compartment call. Check the no-primary control, blocking and detection chemistry before scoring the section (general IHC practice); HPA's Approved rating does not validate this individual slide (HPA tissue IHC). |
| No detectable signal in an otherwise interpretable kidney tubule or thyroid gland section. | Both are reported high-staining cell populations (HPA tissue IHC). First assess tissue preservation, retrieval, antibody dilution and the detection system using suitable controls (general IHC practice); a negative slide alone does not establish absent AHCY. |
💡Expected AHCY appearanceCall a positive result when identifiable cells show nuclear and cytoplasmic signal, especially strong signal in kidney tubules or thyroid glandular cells (HPA tissue IHC); membrane-only colour or staining across empty spaces is suspect (UniProt P23526 topology; general IHC practice).
| Tissue and cell selection | High signal is reported in epididymal and thyroid glandular cells, kidney tubule cells, seminiferous-duct cells and urothelium (HPA tissue IHC). Adipocytes were not detected, making them an unsuitable sole positive control (HPA tissue IHC). |
| Compartment used for scoring | HPA describes tissue staining as nuclear and cytoplasmic, while its supported ICC-IF main location is cytosol (HPA tissue IHC; HPA subcellular ICC-IF). UniProt also lists nucleus, endoplasmic reticulum and melanosome; melanosome identification came from fraction mass spectrometry, not a stated routine IHC pattern (UniProt P23526). |
| IHC antibody evidence | The tissue profile is Approved, with medium staining–RNA consistency; HPA041225 and HPA044675 each have Approved IHC status (HPA tissue IHC; HPA antibodies). These ratings support a reference pattern but do not establish specificity for an untested staining run (general IHC practice). |
| IF/ICC: what pattern is supported? | Cytosolic signal is the supported main ICC-IF location, with images listed for CACO-2, Hep-G2 and KOLF2.1J (HPA subcellular ICC-IF). HPA044675 has Supported ICC status; those IF observations should not be treated as an IHC-P protocol (HPA antibodies). |
| Target-specific preparation limits | The supplied UniProt and HPA records do not report an AHCY-specific fixation effect or antigen-retrieval condition. Select and document IHC-P preparation using validated assay instructions and controls (general IHC practice), without inferring retrieval sensitivity from tissue intensity (HPA tissue IHC). |
| Situation | Likely cause | Next action |
|---|---|---|
| Known high-staining tissue is blank. | An assay-step failure is possible; kidney tubules and thyroid glandular cells are reported high (HPA tissue IHC). | Check section integrity, retrieval, primary-antibody preparation and chromogenic detection against run controls (general IHC practice). |
| Membrane-only colour dominates. | AHCY has no transmembrane segment, and HPA reports nuclear–cytoplasmic tissue staining (UniProt P23526 topology; HPA tissue IHC). | Inspect cellular boundaries and compare a no-primary control; withhold an AHCY call until intracellular staining is credible (general IHC practice). |
| Adipocytes stain strongly. | HPA reports AHCY as not detected in adipocytes; cross-reactivity or endogenous detection activity is possible (HPA tissue IHC; general IHC practice). | Compare no-primary and detection controls, then assess the staining pattern in a reported high-staining cell population (general IHC practice; HPA tissue IHC). |
| Colour coats the whole section. | Nonspecific background or endogenous detection activity may obscure the cellular pattern (general IHC practice). | Review blocking, reagent concentration, washing and no-primary controls before assigning nuclear or cytoplasmic scores (general IHC practice). |
| Only weak signal appears in lung alveolar cells or muscle cells. | HPA reports low signal in alveolar cells, cardiomyocytes and skeletal myocytes (HPA tissue IHC). | Compare with a reported high-staining tissue in the same run before changing assay settings (HPA tissue IHC; general IHC practice). |
| Tissue IHC looks nuclear, but ICC-IF looks mainly cytosolic. | These are the reported observations for the respective applications (HPA tissue IHC; HPA subcellular ICC-IF). | Score each preparation against its own supported reference pattern and controls; investigate only if staining conflicts with that pattern (general IHC practice). |
Experiment evidence
IHC & IF images showing AHCY staining
A04236-1 · IHCIHC analysis of AHCY using anti-AHCY antibody (A04236-1). AHCY was detected in a paraffin-embedded section of human breast cancer tissue. He
A04236-1 · IHCIHC analysis of AHCY using anti-AHCY antibody (A04236-1). AHCY was detected in a paraffin-embedded section of rat liver tissue. Heat mediate
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000101444-AHCY/tissue/cerebral+cortex#imid_10629709
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000101444-AHCY/tissue/colon#imid_10629619
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000101444-AHCY/tissue/liver#imid_10629620
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000101444-AHCY/tissue/kidney#imid_10629637
HPA · ICC-IF CACO-2Human Protein Atlas immunofluorescence, CACO-2, antibody HPA044675; green = AHCY, blue = nucleus, red = microtubules.
HPA · ICC-IF CACO-2Human Protein Atlas immunofluorescence, CACO-2, antibody HPA044675; green = AHCY, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for AHCY IHC & IF
🧪Run kidney first: tubular cells should stain strongly (HPA: High in cells in tubules). Use adipose tissue as the negative tissue (HPA: Not detected in adipocytes); on the kidney slide, cells without specific staining should show only counterstain or background, but no particular kidney cell type is established as AHCY-negative by the supplied HPA rows.
Positive control tissue: Epididymis (Glandular cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show AHCY in CACO-2, Hep-G2, KOLF2.1J, with annotated localisation: Cytosol (supported) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control and a matched nonimmune rabbit IgG isotype control (selected-SKU IHC caption: rabbit primary antibody; standard IHC practice). Confirm specificity with AHCY-knockout tissue if available, quench endogenous peroxidase before HRP/DAB detection, and block endogenous biotin if using a biotin-based system (standard IHC practice).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected-SKU paraffin-section caption does not state a fixative (selected-SKU IHC caption). Heat retrieval in EDTA at pH 8.0 was used, but whether AHCY staining depends on retrieval is unreported (selected-SKU IHC caption). Frozen sections are not established as easier; HPA ICC-IF supports cytosolic localisation but does not establish tissue IF performance, and kidney sections need background assessment when using HRP/DAB (HPA: cytosol supported; standard IHC practice).
HPA tissue IHC evidence for AHCY
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — 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 |
|---|---|---|---|---|
| Epididymis | Glandular cells | High | Protein (IHC) | HPA → |
| Kidney | Cells in tubules | High | Protein (IHC) | HPA → |
| Testis | Cells in seminiferous ducts | High | Protein (IHC) | HPA → |
| Thyroid gland | Glandular cells | High | Protein (IHC) | HPA → |
| Urinary bladder | Urothelial cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced AHCY IHC Tips
Troubleshoot AHCY staining in paraffin section chromogenic IHC using the selected antibody’s tissue image and independent expression evidence.
What retrieval should I use when AHCY staining is weak in paraffin sections?
Start with heat-mediated antigen retrieval in EDTA at pH 8.0 for paraffin sections (datasheet A04236-1). The selected tissue image used this retrieval before 2 μg/ml primary antibody overnight at 4°C, so reproduce those conditions when troubleshooting weak DAB signal (caption A04236-1). Compare matched sections processed with and without retrieval while holding section thickness, detection chemistry, and development time constant (standard IHC practice). If signal remains weak, adjust heating duration within limits that preserve tissue morphology; excessive heating can damage sections or increase background (standard IHC practice).
Could fixation explain inconsistent AHCY staining between specimens?
The selected caption describes a paraffin-embedded human breast cancer section but does not state its fixative, so AHCY-specific fixation sensitivity is unknown (caption A04236-1). Record the fixative, fixation duration, and processing history for each specimen before attributing a staining difference to fixation (standard IHC practice). For a controlled comparison, process matched sections together, apply EDTA retrieval at pH 8.0, and include the same positive control (datasheet A04236-1; standard IHC practice). Assess morphology and staining together, then test any proposed fixation change on matched material before concluding that an AHCY epitope was lost (standard IHC practice).
How should I assess nuclear versus cytoplasmic AHCY staining?
Expect a substantial cytoplasmic component: the HPA subcellular assessment supports cytosolic localisation, while its tissue IHC profile describes nuclear and cytoplasmic expression (HPA subcellular; HPA tissue IHC). UniProt also lists the nucleus, endoplasmic reticulum, and melanosome, with melanosome identification based on fraction mass spectrometry (UniProt P23526 localisation). Score nuclear and cytoplasmic DAB separately in morphologically intact cells, using the same compartment definitions across sections (standard IHC practice). Treat a compartment seen only at tissue edges, in damaged areas, or with mismatched control staining as uncertain until independently reproduced (standard IHC practice).
Can staining distinguish AHCY isoforms or modified forms?
AHCY has 2 annotated isoforms, but the supplied antibody caption does not identify its epitope or establish isoform selectivity (UniProt P23526 isoforms; caption A04236-1). The record lists modified residues, including phosphoserine 183 and phosphotyrosine 193, without evidence that these changes alter this antibody’s tissue staining (UniProt P23526 modified residues). AHCY has no annotated transmembrane segment or glycosylation sites, which describes the protein but does not locate the antibody’s epitope (UniProt P23526 topology and glycosylation). Interpret chromogenic signal as AHCY immunoreactivity; assign an isoform or modification only after validation with an epitope-defined reagent or another independent method (standard IHC practice).
How can I cross-check the IHC pattern with multiplex IF?
For an IF cross-check, pair AHCY with a marker that identifies the cell population being scored, such as the tubular cells used as an HPA positive reference (HPA tissue IHC; standard IF practice). Choose spectrally separated fluorophores, favouring a longer wavelength channel for weak AHCY signal when tissue autofluorescence interferes, and inspect single-channel controls (standard IF practice). Because AHCY is intracellular and has no transmembrane segment, permeabilise sufficiently to access its cytosolic and potential nuclear epitopes without assuming a membrane-facing epitope (UniProt P23526 topology and localisation; standard IF practice). Compare compartment patterns with the supported cytosolic IF localisation, while keeping the chromogenic IHC and IF readouts separately controlled (HPA subcellular; standard IF practice).
What should I check when DAB background obscures AHCY staining?
The selected tissue image used 10% goat serum before primary antibody and an HRP-based DAB readout (caption A04236-1). Check a no-primary section, endogenous peroxidase blocking, and the timing of DAB development to identify background from detection chemistry (standard IHC practice). Keep blocking and development conditions consistent while comparing the reported 2 μg/ml primary incubation overnight at 4°C with a lower antibody concentration if diffuse staining persists (caption A04236-1; standard IHC practice). Evaluate granular or brown material against unstained and no-primary sections before treating it as intracellular AHCY, particularly where tissue pigment or damaged cells complicate interpretation (standard IHC practice).
How should I quantify AHCY across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and score viable cells separately from damaged or necrotic regions before measuring chromogenic AHCY signal (standard IHC practice). An H-score multiplies the percentage of cells at each intensity grade by that grade, using 0–3 intensity grades and a possible 0–300 range (standard IHC practice). Record nuclear and cytoplasmic scores separately because both compartments appear in the reported tissue profile, and retain percent positive cells alongside intensity (HPA tissue IHC; standard IHC practice). Normalise comparisons to the same cell population, evaluated area, staining batch, and positive control; for counts, report positive cells per mm² of viable tissue (standard IHC practice).
When is an apparent AHCY-positive cell likely an artefact?
A reproducible intracellular signal in intact cells is more credible when its cytoplasmic component agrees with supported cytosolic localisation and matched controls behave as expected (HPA subcellular; standard IHC practice). HPA reports high staining in kidney tubular cells and no detected staining in adipocytes, providing cell-specific references rather than a universal cutoff (HPA tissue IHC). Question signal confined to section edges, necrosis, or no-primary controls, and investigate endogenous peroxidase activity before accepting brown DAB deposits as AHCY (standard IHC practice). Nuclear staining can be plausible in tissue, but a new compartment or unexpected cell population needs repeat staining and independent validation before biological interpretation (HPA tissue IHC; standard IHC practice).
Boster reagents
Best AHCY / Adenosylhomocysteinase IHC Antibodies
The catalog antibody has IHC data from paraffin sections of human breast cancer and rat liver (IHC image captions); human, mouse and rat reactivity is listed (catalog).
Real IHC data
Anti-AHCY Antibody ®
Cat # A04236-1
A04236-1 is listed for IHC and has images from paraffin sections of human breast cancer and rat liver (catalog applications; IHC image captions). Its listed reactivity is human, mouse and rat, while its pictured IHC examples cover human and rat tissue (catalog reactivity; IHC image captions).
Which to pick: Choose A04236-1 for paraffin-section IHC: its own captions show staining at 2 μg/ml after EDTA pH 8.0 retrieval; the fixative is unreported (IHC image captions). For IF/ICC, this SKU has no listed IF/ICC application or IF image, so the payload provides no IF/ICC validation (catalog applications; IF image alts). For cross-species work, A04236-1 lists human, mouse and rat reactivity, but its pictured IHC evidence covers human and rat only (catalog reactivity; IHC image captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P23526 (SAHH_HUMAN, Adenosylhomocysteinase).
- Human Protein Atlas. AHCY tissue IHC expression (reliability: Approved).
- Human Protein Atlas. AHCY subcellular location (ICC-IF): Localized to the cytosol..
- Human Protein Atlas. AHCY antibody validation summary (2 antibodies).
- Screening and cellular validation of prognostic genes regulated by super enhancers in oral squamous cell carcinoma. Bioengineered 2021 — PMC8810015.
- Wumei Pill Ameliorates AOM/DSS-Induced Colitis-Associated Colon Cancer through Inhibition of Inflammation and Oxidative Stress by Regulating S-Adenosylhomocysteine Hydrolase- (AHCY-) Mediated Hedgehog Signaling in Mice. Oxidative medicine and cellular longevity 2022 — PMC9345734.
- Integrated bulk and single-cell transcriptomics unveil the role of AHCY in cell cycle regulation of endometrial cancer. Frontiers in oncology 2026 — PMC13538023.
- Multi-omics characterization identifies AHCY as a prognostic biomarker driving immunometabolic reprogramming in bladder cancer. Translational oncology 2026 — PMC12813242.
- PubMed PMID:2596825 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
- PubMed PMID:11780052 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for AHCY (Adenosylhomocysteinase). Expected localisation Nuclear and cytoplasmic staining (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A04236-1). Validated IHC antibody: A04236-1 at 2-5 μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Epididymis (Glandular cells); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human breast cancer tissue; fixative not specified (datasheet A04236-1); 2. Fixation: Image fixative and duration unreported (datasheet A04236-1); 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 retrieval: EDTA pH 8.0 (datasheet A04236-1); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A04236-1); 8. Primary antibody: Rabbit anti-AHCY, 2-5 μg/ml (datasheet A04236-1); 9. Primary incubation: Overnight at 4 °C (datasheet A04236-1). Watch for: Fixation: Keep fixation consistent across sections. (standard IHC practice; not target-specific); Caveat: Staining and RNA show medium consistency (HPA tissue IHC); Isoform / epitope: 2 isoforms; epitope coverage is unknown (UniProt); Regulation: Specific regulators unreported (UniProt). Evidence: UniProt P23526, HPA tissue IHC · Approved, HPA ICC-IF · cytosol., Datasheet A04236-1, PMC9345734. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/AHCY/immunohistochemistry-ihc-guide.