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FZD5 / Frizzled-5 · IHC design guide
Design Immunohistochemistry for FZD5
Plan FZD5 IHC in paraffin sections using the cytoplasmic tissue pattern reported for gastrointestinal glands, placenta and brain (HPA tissue IHC). Compare that pattern with its annotated membrane and Golgi locations when interpreting staining (UniProt).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
FZD5 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 in tissue; membrane and Golgi annotated (HPA tissue IHC; UniProt) | |
| Staining pattern | Cytoplasmic in GI glands, trophoblasts and neurons (HPA tissue IHC) | |
| Antigen retrieval | Tris-EDTA pH 9.0 HIER, heat-mediated (datasheet A04292-1) | |
| Positive control | Caudate+4 more · see all | |
| Negative control | Adipose tissue+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A04292-1) | |
| Caveat | Tissue IHC is cytoplasmic despite membrane annotation (HPA tissue IHC; UniProt) | |
| Regulation | Expression regulation is not annotated (UniProt) | |
| Isoform / epitope | No isoforms listed; epitope location depends on topology (UniProt) |
Section 1
Recommended FZD5 IHC & IF Protocols
The catalog antibody has a heat-mediated, pH 9.0 IHC-P protocol (datasheet A04292-1). Four published FZD5 IHC protocols provide tissue-specific starting points (PMC7717951; PMC6487103; PMC6007469; PMC13560161).
| Sample | Paraffin-embedded human tonsil tissue; fixative not specified (datasheet A04292-1) |
| Fixation | Image fixative and duration unreported (datasheet A04292-1); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat retrieval: Tris-EDTA pH 9.0 (datasheet A04292-1); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% normal serum of the secondary host, 30 min, room temperature (standard) |
| Primary antibody | Rabbit anti-FZD5, 1:100-1:300 (datasheet A04292-1) |
| 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 | FZD5-positive staining in neuronal cells of caudate (HPA tissue IHC: Medium). HPA tissue profile: Cytoplasmic expression mainly in glandular cells in the gastrointestinal tract, placenta and the brain. No signal in the no-primary control. |
💡Decision noteStart with Tris-EDTA pH 9.0 for the catalog antibody (datasheet A04292-1). For renal sections, citrate pH 6.0 is a published alternative (PMC7717951).
Section 2
What Is the Expected FZD5 Staining Pattern?
FZD5 is a 7-transmembrane receptor at the plasma membrane and Golgi apparatus (UniProt Q13467 topology; UniProt Q13467 subcellular location). In paraffin-section IHC, expect staining in glandular cells of the gastrointestinal tract, trophoblastic cells of the placenta, and selected brain cells; HPA describes the observed pattern as mainly cytoplasmic (HPA tissue IHC). HPA rates its tissue IHC evidence Approved, pending external verification (HPA tissue IHC).
| Cell-associated chromogenic staining in duodenal or small-intestinal glandular cells, placental trophoblastic cells, or selected neurons (HPA tissue IHC). | This fits HPA's Medium staining in those cell populations (HPA tissue IHC). Membrane-associated and cytoplasmic signal can both be plausible: UniProt places FZD5 at the plasma membrane and Golgi, while HPA describes mainly cytoplasmic tissue staining (UniProt Q13467 subcellular location; HPA tissue IHC). |
| Predominantly nuclear staining, with little convincing membrane-associated or cytoplasmic signal. | A nuclear-dominant pattern does not match the reported FZD5 locations or HPA tissue pattern (UniProt Q13467 subcellular location; HPA tissue IHC). Treat it as a possible staining artefact and compare it with a negative reagent control before scoring it as FZD5 (general IHC practice). |
| Strong staining in adipocytes or cardiomyocytes, while expected positive cells show little signal. | HPA reports FZD5 as Not detected in those specific cell populations (HPA tissue IHC). Consider antibody cross-reactivity or endogenous chromogenic activity; inspect cellular boundaries and control sections before assigning the signal to FZD5 (general IHC practice). |
| Diffuse color across cells, stroma, and empty slide areas, without a consistent cellular pattern. | This does not resemble HPA's cell-restricted observations (HPA tissue IHC). Background from nonspecific antibody binding, detection reagents, or incomplete washing can obscure a real signal; the pattern alone cannot identify which step caused it (general IHC practice). |
| No convincing staining in duodenal glandular cells or placental trophoblastic cells. | Both populations have Medium HPA tissue IHC staining (HPA tissue IHC). A blank result warrants a check of section integrity, retrieval, antibody and detection performance before concluding that the specimen lacks FZD5 (general IHC practice). |
💡Expected FZD5 appearanceCall a result positive when moderate, cell-associated membrane-associated or cytoplasmic staining occurs in an HPA-positive cell population; predominant nuclear staining or diffuse cell-independent color is suspect (HPA tissue IHC; UniProt Q13467 subcellular location; general IHC practice).
| Compartment and topology | FZD5 has 7 transmembrane segments and is reported at the plasma membrane and Golgi (UniProt Q13467 topology; UniProt Q13467 subcellular location). HPA's paraffin-section profile is mainly cytoplasmic, so a cytoplasmic result is compatible with its observations; topology alone cannot establish the appearance of this antibody's IHC stain (HPA tissue IHC). |
| Choice of comparison cells | HPA reports Medium staining in duodenal and small-intestinal glandular cells and placental trophoblastic cells, but Not detected in adipocytes and cardiomyocytes (HPA tissue IHC). Compare named cell populations, rather than treating an entire organ as uniformly positive or negative (HPA tissue IHC). |
| Strength of pattern evidence | The tissue profile and listed IHC antibody HPA052361 are Approved; the profile is pending external verification, and the antibody is not listed as Enhanced (HPA tissue IHC; HPA antibodies). Use the reported pattern as a reference, while treating an unexpected result as something to investigate rather than an automatic biological finding. |
| Extracellular region and processing | UniProt places an extracellular region at residues 27–238, lists N-glycosylation sites at 47 and 151, and describes a signal peptide at 1–26 (UniProt Q13467 topology; UniProt Q13467 processing; UniProt Q13467 glycosylation). The supplied evidence gives no antibody epitope, so these annotations do not predict retrieval needs or staining loss. |
| Antigen retrieval | Retrieval is a routine variable to check when paraffin-section IHC lacks signal (general IHC practice). The supplied UniProt and HPA records give no FZD5-specific retrieval condition or fixation-sensitivity finding; select and assess conditions using the IHC-validated antibody's instructions and controls. |
| IF/ICC Q&A: should the IHC pattern be expected in cells? | HPA summarizes FZD5's ICC-IF location as Membrane but provides no main location or cell-line images in this payload (HPA subcellular). UniProt also reports plasma membrane and Golgi locations (UniProt Q13467 subcellular location). These records support a location check, not a validated IF/ICC staining appearance or protocol. |
| Situation | Likely cause | Next action |
|---|---|---|
| Known-positive glandular or trophoblastic cells are blank (HPA tissue IHC). | The staining run may have failed, or the specimen may have inadequate evaluable cells; the image alone cannot distinguish these possibilities (general IHC practice). | Check section morphology, the run's positive control, retrieval record, primary-antibody application, and detection reagents; repeat with a documented working run if a control failed (general IHC practice). |
| Signal is weak in an HPA-positive cell population (HPA tissue IHC). | Low assay sensitivity or an unsuitable retrieval or antibody condition may reduce visible chromogen (general IHC practice). HPA's Medium category does not specify a required intensity for every specimen (HPA tissue IHC). | Compare cells on the same run with a positive control and the documented IHC conditions; adjust one routine assay variable at a time if controls support troubleshooting (general IHC practice). |
| Color appears in adipocytes or cardiomyocytes, where HPA reports Not detected (HPA tissue IHC). | Cross-reactive binding or endogenous detection activity is possible; HPA's observation does not identify which explanation applies (HPA tissue IHC; general IHC practice). | Review a negative reagent control and an expected positive cell population, then check endogenous-enzyme blocking and detection controls for the chromogenic method used (general IHC practice). |
| Predominantly nuclear staining replaces the expected cell-associated pattern. | The location conflicts with UniProt's membrane and Golgi annotation and HPA's mainly cytoplasmic profile (UniProt Q13467 subcellular location; HPA tissue IHC). Nonspecific staining is possible (general IHC practice). | Check whether the color persists in a negative reagent control, inspect counterstain separately, and avoid scoring nuclear-only signal as FZD5 without independent support (general IHC practice). |
| Diffuse chromogen makes individual positive cells hard to identify. | Nonspecific binding, residual detection activity, or insufficient washing can raise background in chromogenic IHC (general IHC practice). | Compare a negative reagent control, confirm the blocking and wash steps used, and assess whether recognizable HPA-positive cells remain above background (general IHC practice; HPA tissue IHC). |
| A section from an HPA-positive tissue disagrees with the reference pattern (HPA tissue IHC). | The sampled cell population, tissue condition, or assay performance may differ; HPA's Approved profile is pending external verification (HPA tissue IHC; general IHC practice). | Target-specific fixation effects are not established by the supplied assay evidence. Verify with a matched IHC source before attributing a result to fixation. |
Experiment evidence
IHC & IF images showing FZD5 staining
A04292-1 · IHCImmunohistochemical analysis of paraffin-embedded human tonsil. 1, Antibody was diluted at 1:200 (4° overnight). 2, Tris-EDTA, pH9.0 was use
A04292-1 · IFImmunofluorescence analysis of A549 cells, using FZD5 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/ENSG00000163251-FZD5/tissue/cerebral+cortex#imid_15687423
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000163251-FZD5/tissue/colon#imid_15687365
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000163251-FZD5/tissue/liver#imid_15687350
HPA · PancreasHuman Protein Atlas IHC, Pancreas. Source: https://www.proteinatlas.org/ENSG00000163251-FZD5/tissue/pancreas#imid_15687309
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for FZD5 IHC & IF
🧪Run cerebellum first and look for staining in Purkinje cells (HPA: Medium in cerebellar Purkinje cells). Run adipose tissue as the negative tissue (HPA: Not detected in adipocytes); on the cerebellum slide, use cells that remain at counterstain/background level as internal negatives, without assuming every other cell type is negative.
Positive control tissue: Caudate (Neuronal cells, HPA Medium)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA carries no ICC-IF cell line for FZD5; derive a cell-line control from the positive tissue's cell type (Neuronal cells) and confirm it by RNA or western blot first.
Technical controls: Include a no-primary (secondary-only) slide, a host-species- and clonality-matched isotype control, and FZD5 knockout tissue or a peptide-block control if the immunizing peptide is available (standard IHC controls). Quench endogenous peroxidase for chromogenic detection and check Purkinje-cell pigment against the controls when scoring (standard IHC practice).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected A04292-1 paraffin-section caption does not report a fixative (selected-SKU caption). That caption uses Tris-EDTA retrieval at pH 9.0, but does not establish whether retrieval is required; frozen sections and IF cannot be ranked as easier from the supplied evidence (selected-SKU caption; HPA: no ICC-IF cell-line images). Interpret any signal against the expected membrane and Golgi localization and check neuronal pigment before scoring chromogenic staining (UniProt Q13467 localization; standard IHC practice).
HPA tissue IHC evidence for FZD5
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Pending external verification.). 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 |
|---|---|---|---|---|
| Caudate | Neuronal cells | Medium | Protein (IHC) | HPA → |
| Cerebellum | Purkinje cells | Medium | Protein (IHC) | HPA → |
| Duodenum | Glandular cells | Medium | Protein (IHC) | HPA → |
| Hippocampus | Neuronal cells | Medium | Protein (IHC) | HPA → |
| Placenta | Trophoblastic cells | Medium | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Appendix | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Breast | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Colon | Endothelial cells | Not detected | Protein (IHC) | HPA → |
| Esophagus | Squamous epithelial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced FZD5 IHC Tips
Troubleshoot FZD5 staining in paraffin sections by checking retrieval, cell type, and compartment before comparing signal intensity across samples.
Where should I start if FZD5 staining is weak after antigen retrieval?
Start paraffin-section IHC with heat-mediated Tris-EDTA at pH 9.0 (datasheet A04292-1). The catalog antibody's human tonsil image used that retrieval, followed by primary antibody at 1:200 overnight at 4°C (datasheet A04292-1). If staining is weak, compare retrieval heating and cooling conditions on matched sections while keeping antibody dilution and detection constant (standard IHC practice). Include a section processed without primary antibody to distinguish weak specific signal from detection background (standard IHC practice). Record the heating duration and tissue integrity for each condition; excessive retrieval can damage morphology and complicate scoring (standard IHC practice).
How should I investigate whether fixation is affecting FZD5 IHC?
Target-specific fixation sensitivity is unknown: the catalog image identifies paraffin-embedded tonsil but does not state its fixative (datasheet A04292-1). Record the fixative, fixation duration in hours, and processing history for every section before comparing staining (standard IHC practice). On matched material, vary one fixation or processing condition at a time and keep Tris-EDTA retrieval at pH 9.0 constant (datasheet A04292-1; standard IHC practice). Evaluate tissue morphology alongside staining because processing differences can alter apparent intensity and compartment boundaries (standard IHC practice). Do not attribute a weak result to FZD5-specific fixation sensitivity without a direct comparison (standard IHC practice).
What staining pattern is plausible for FZD5 in paraffin sections?
Assess membrane-associated staining first: FZD5 localizes to the plasma membrane and Golgi apparatus membrane (UniProt Q13467 localization). Its 7 transmembrane segments make a purely nuclear pattern difficult to reconcile with the annotated protein (UniProt Q13467 topology). Cytoplasmic staining can still be plausible because the tissue atlas reports it mainly in gastrointestinal glandular cells, placenta, and brain (HPA tissue IHC). Review intact cell boundaries and perinuclear signal against the counterstain before assigning a compartment (standard IHC practice). Compare glandular or neuronal cells with adjacent structures on the same slide, since tissue context helps distinguish cellular staining from deposited chromogen (HPA tissue IHC; standard IHC practice).
How can epitope position change my interpretation of FZD5 staining?
Check the antibody's documented immunogen before treating a staining pattern as epitope-specific; none is provided in this payload (standard IHC practice). FZD5 has an extracellular region spanning residues 27–238 and a cytoplasmic tail at 522–585 (UniProt Q13467 topology). Its signal peptide spans 1–26, while the annotated mature chain begins at 27 (UniProt Q13467 processing). Glycosylation sites at 47 and 151 provide further context for an extracellular epitope, but do not establish antibody recognition (UniProt Q13467 glycosylation; standard IHC practice). No isoforms are annotated in the supplied record, so do not assign staining differences to a specific isoform (UniProt Q13467 isoforms).
How should I extend this FZD5 IHC assessment to multiplex IF?
Treat IF as a separate validation: the supplied antibody image documents paraffin-section chromogenic IHC, and the HPA payload lists no ICC/IF images (datasheet A04292-1; HPA subcellular). Pair FZD5 with a validated marker for the expected glandular, neuronal, or trophoblastic cell type and inspect each channel separately (HPA tissue IHC; standard IF practice). Select fluorophores after measuring tissue autofluorescence, and include single-label and no-primary controls to assess bleed-through and background (standard IF practice). Choose permeabilisation according to the documented epitope: access to the cytoplasmic tail differs from access to an extracellular region (UniProt Q13467 topology; standard IF practice). Do not transfer the IHC antibody dilution of 1:200 to IF without titration (datasheet A04292-1; standard IF practice).
How can I reduce diffuse or patchy FZD5 chromogenic background?
First compare the stained section with a no-primary control and check whether chromogen follows cells or spreads across tissue (standard IHC practice). Use a peroxidase block for peroxidase-based detection, then check whether residual signal persists without primary antibody (standard IHC practice). The catalog tonsil image used primary antibody at 1:200 overnight at 4°C and secondary antibody at 1:200 for 45 minutes at room temperature (datasheet A04292-1). If background dominates, titrate antibody concentrations and washing while keeping retrieval at pH 9.0 fixed during that comparison (datasheet A04292-1; standard IHC practice). Examine section edges and damaged areas separately from intact cells before calling diffuse color FZD5-positive (standard IHC practice).
What should I score when comparing FZD5 IHC across sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the eligible cell population and compartment before scoring, using the same rules for every section (standard IHC practice). For glandular, neuronal, or trophoblastic populations, report the percentage of positive eligible cells and an intensity-based H-score when staining permits reliable cell boundaries (HPA tissue IHC; standard IHC practice). Calculate H-score as 0–300 from percentages at intensities 0–3, and specify whether membrane-associated and cytoplasmic signal were scored separately (standard IHC practice; UniProt Q13467 localization). Normalize counts to the number of eligible cells, or area-based density to analyzed tissue area in mm² (standard IHC practice). Keep retrieval, imaging, counterstain, and positivity thresholds consistent across samples (standard IHC practice).
How do I distinguish credible FZD5 positivity from staining artefacts?
A credible result follows intact cells in a plausible compartment: FZD5 is annotated at the plasma membrane and Golgi membrane (UniProt Q13467 localization). Compare the stained cell type with the HPA pattern, including gastrointestinal glandular cells, trophoblastic cells, and neuronal cells, without treating that pattern as independent antibody validation (HPA tissue IHC). Uniform nuclear color, section-edge staining, and deposits over necrotic regions warrant review against morphology and a no-primary control (UniProt Q13467 localization; standard IHC practice). Residual signal after omitting primary antibody can indicate endogenous enzyme activity or detection background in a peroxidase-based assay (standard IHC practice). Interpret intensity alongside compartment, cell identity, and controls rather than calling every brown area FZD5-positive (standard IHC practice).
Boster reagents
Best FZD5 / Frizzled-5 IHC Antibodies
Anti-FZD5 options list human, mouse and rat reactivity (catalog: reactivity); A04292-1 has human tonsil IHC and A549-cell IF images (A04292-1 image captions).
Real IHC data
Anti-Frizzled-5 FZD5 Antibody
Cat # A04292-1
A04292-1 lists IHC and IF, with images of paraffin-embedded human tonsil and A549 cells (A04292-1 applications; image captions). A04292-2 lists IHC, IF and ICC, while A04292Y500 lists IHC and IF; both list human, mouse and rat reactivity, but neither has an IHC image in this payload (catalog: applications, reactivity, IHC image alts).
Which to pick: For tissue IHC, choose A04292-1: its own caption documents paraffin-embedded human tonsil stained at 1:200 after Tris-EDTA retrieval at pH 9.0; the fixative is unreported (A04292-1 IHC caption). For IF/ICC, A04292-2 lists both applications and is polyclonal (A04292-2 applications; dilution_raw); A04292-1 also has an A549-cell IF image (A04292-1 IF caption). For cross-species IHC, start with A04292-1 because it lists human, mouse and rat reactivity, while recognising that its IHC image documents human tissue only (A04292-1 reactivity; IHC caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q13467 (FZD5_HUMAN, Frizzled-5).
- Human Protein Atlas. FZD5 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. FZD5 subcellular location (ICC-IF): Membrane.
- Human Protein Atlas. FZD5 antibody validation summary (1 antibodies).
- Comparative Assessment of the WNT/β-Catenin Pathway, CacyBP/SIP, and the Immunoproteasome Subunit LMP7 in Various Histological Types of Renal Cell Carcinoma. Frontiers in oncology 2020 — PMC7717951.
- Wingless-Type MMTV Integration Site Family Member 5a Is a Key Secreted Islet Stellate Cell-Derived Product that Regulates Islet Function. International journal of endocrinology 2019 — PMC6487103.
- Role of WNT5A receptors FZD5 and RYK in prostate cancer cells. Oncotarget 2018 — PMC6007469.
- FZD5 drives macrophage-mediated immunomodulation and predicts prognosis in glioma: evidence from single-cell sequencing. BMC cancer 2026 — PMC13560161.
- PubMed PMID:8626800 — UniProt-cited evidence.
- PubMed PMID:11408929 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for FZD5 (Frizzled-5). Expected localisation Cytoplasmic in tissue; membrane and Golgi annotated (HPA tissue IHC; UniProt). Antigen retrieval: Tris-EDTA pH 9.0 HIER, heat-mediated (datasheet A04292-1). Validated IHC antibody: A04292-1 at 1:100-1:300 (the product's own IHC validation image is shown). Controls (HPA): positive Caudate (Neuronal cells); negative Adipose tissue. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Paraffin-embedded human tonsil tissue; fixative not specified (datasheet A04292-1); 2. Fixation: Image fixative and duration unreported (datasheet A04292-1); 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: Tris-EDTA pH 9.0 (datasheet A04292-1); 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-FZD5, 1:100-1:300 (datasheet A04292-1); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A04292-1); Caveat: Tissue IHC is cytoplasmic despite membrane annotation (HPA tissue IHC; UniProt); Isoform / epitope: No isoforms listed; epitope location depends on topology (UniProt); Regulation: Expression regulation is not annotated (UniProt). Evidence: UniProt Q13467, HPA tissue IHC · Approved, HPA ICC-IF · Membrane, Datasheet A04292-1, PMC7717951, PMC6487103. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/FZD5/immunohistochemistry-ihc-guide.