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IL9R / Interleukin-9 receptor · IHC design guide
Design Immunohistochemistry for IL9R
Plan chromogenic IL9R IHC in paraffin sections using the membrane topology and tissue staining evidence (UniProt; HPA tissue IHC). The guide highlights high staining in round or early spermatids, lower staining in several other cell populations, and a reported off-target binding concern (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
IL9R 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; secreted form also annotated (UniProt) | |
| Staining pattern | Subset of immune cells; compartment unreported (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Testis | |
| 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). Selected-image fixative and duration unreported (datasheet A07490-1); verify before use. | |
| Caveat | Presumed off-target binding may mimic IL9R staining (HPA tissue IHC) | |
| Regulation | Expression regulation unreported (UniProt) | |
| Isoform / epitope | 3 isoforms; check extracellular versus cytoplasmic epitope (UniProt) |
Section 1
Recommended IL9R IHC & IF Protocols
The catalog antibody protocol is paired with a published IL9R IHC protocol for paraffin-embedded skin sections (PMC4967550).
| Sample | Paraffin-embedded human liver carcinoma tissue; fixative not specified (datasheet A07490-1) |
| Fixation | Image fixative and duration unreported (datasheet A07490-1); 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-IL9R, 1:50-1:200 (datasheet A07490-1) |
| 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 | IL9R-positive staining in round or early spermatids of testis (HPA tissue IHC: High). HPA tissue profile: Expression in a subset of immune cells. No signal in the no-primary control. |
💡Decision noteStart with citrate pH 6.0 retrieval at 95–98 °C for 20 min (page retrieval); compare TRIS-EDTA pH 9 when adapting the skin protocol (PMC4967550).
Section 2
What Is the Expected IL9R Staining Pattern?
IL9R is a membrane receptor with an extracellular region and cytoplasmic tail (UniProt Q01113 topology); UniProt also annotates a secreted location (UniProt Q01113 subcellular). In paraffin tissue IHC, look first for staining in round or early spermatids, reported as high, and a subset of immune cells (HPA tissue IHC). HPA rates the tissue profile Approved, pending external verification, and notes presumed off-target binding that was disregarded (HPA tissue IHC).
| Distinct staining in round or early spermatids, especially near cell outlines. | This fits the high testis signal reported by HPA (HPA tissue IHC). A membrane-associated appearance is consistent with receptor topology (UniProt Q01113 topology), but HPA's tissue record does not establish the exact stained compartment in these cells. |
| Predominantly nuclear staining, with little staining near cell outlines. | A nuclear-only pattern does not fit the membrane topology or HPA membrane summary (UniProt Q01113 topology; HPA subcellular). Check the counterstain and detection controls before treating nuclear color as IL9R. |
| Strong staining in nearly every cell of a tissue reported as negative. | That conflicts with HPA's cell-specific observations; adipocytes in adipose tissue and respiratory epithelial cells in bronchus are examples reported as not detected (HPA tissue IHC). Cross-reactivity or endogenous chromogenic activity is plausible; neither can be diagnosed from appearance alone. |
| Diffuse color across cells, stroma, and slide background. | This does not resolve into the cell-specific profile described by HPA (HPA tissue IHC). As a general IHC check, inspect reagent-only and detection controls, washing, blocking, and antibody concentration before scoring positive cells. |
| No visible staining in round or early spermatids on an otherwise interpretable section. | That misses the strongest supplied positive reference (HPA tissue IHC: high in round or early spermatids). Review section quality, retrieval and detection controls as general IHC checks; the supplied sources give no IL9R-specific fixation sensitivity or working dilution. |
💡Expected IL9R appearanceCall a result convincing when round or early spermatids show distinct, high cell-associated staining (HPA tissue IHC), with membrane-associated localization biologically plausible (UniProt Q01113 topology); isolated nuclear color or uniform staining of reported negative cell types is suspect (HPA subcellular; HPA tissue IHC).
| Receptor topology and epitope location | IL9R has extracellular residues 41–270, a transmembrane segment at 271–291, and a cytoplasmic region at 292–521 (UniProt Q01113 topology). These positions guide interpretation of a documented antibody epitope; no epitope for the catalog antibody is supplied here. |
| Processing and secreted annotation | The signal peptide spans residues 1–40, and the annotated chain begins at residue 41 (UniProt Q01113 processing). UniProt also lists IL9R as secreted (UniProt Q01113 subcellular); these facts alone cannot establish that diffuse tissue staining is a shed receptor. |
| Glycosylation and isoforms | UniProt lists glycosylation sites at residues 117 and 156 and 3 isoforms (UniProt Q01113). Their effect on this antibody's paraffin-section staining is not supplied, so do not assign a staining difference to either feature without antibody-specific evidence. |
| Strength and scope of tissue evidence | HPA reports high staining in round or early spermatids, low staining in macrophages and selected glandular or urothelial cells, and no detection in listed negative cells (HPA tissue IHC). Its Approved rating is pending external verification, with presumed off-target binding disregarded (HPA tissue IHC). |
| Antibody validation and IF evidence | Both listed antibodies, HPA063240 and HPA064557, have IHC Approved status; neither has an ICC status in the supplied record (HPA antibodies). HPA summarizes the subcellular location as membrane but supplies no ICC-IF image cell lines (HPA subcellular). |
| Situation | Likely cause | Next action |
|---|---|---|
| The testis positive reference is blank. | The result conflicts with HPA's high signal in round or early spermatids (HPA tissue IHC); a failed stain or unsuitable section remains possible. | Confirm the expected cells are present, then review retrieval, primary-antibody application and chromogenic detection with routine positive and reagent controls. No source here identifies an IL9R-specific retrieval condition. |
| Staining looks mainly nuclear. | A nuclear-only result is discordant with membrane topology (UniProt Q01113 topology; HPA subcellular); nonspecific color or counterstain overlap may be involved. | Compare with the no-primary control and inspect cell boundaries at higher magnification. Score only resolved cell staining, and do not infer a new nuclear location from this section alone. |
| Reported negative cell types stain broadly. | Uniform color in adipocytes or bronchial respiratory epithelium conflicts with HPA's not-detected entries (HPA tissue IHC). Cross-reactivity or endogenous enzyme activity may contribute. | Check a no-primary detection control and the routine endogenous-activity block for the chosen chromogen. Compare the distribution with a reported positive cell population (HPA tissue IHC). |
| Background obscures individual cells. | Diffuse background cannot be matched reliably to HPA's subset-of-immune-cells profile (HPA tissue IHC). In routine IHC, excessive antibody, insufficient blocking or incomplete washing can contribute. | Use routine negative controls, verify blocking and washing, and titrate the antibody against a positive reference. The supplied record provides no catalog dilution or IL9R-specific adjustment. |
| Low-level lung or gut staining seems weaker than testis. | HPA reports low macrophage staining in lung and low glandular staining in duodenum, small intestine, colon and rectum, versus high staining in round or early spermatids (HPA tissue IHC). | Score the named cell populations separately and compare runs with the same positive reference. Avoid calling an entire organ positive or negative from a small, low-signal cell population. |
| Can this IHC pattern be used as an IF/ICC protocol? | HPA's membrane summary provides no ICC-IF image cell lines, and the listed antibodies have no ICC status in this record (HPA subcellular; HPA antibodies). | Treat membrane localization as an interpretation cue (UniProt Q01113 topology; HPA subcellular). Use the separate IF/ICC guide for method design; do not transfer an IHC-P condition as a validated IF setting. |
Experiment evidence
IHC & IF images showing IL9R staining
A07490-1 · IHCImmunohistochemistry (IHC) analyzes of IL-9R (L514) pAb in paraffin-embedded human liver carcinoma tissue at 1:100.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000124334-IL9R/tissue/cerebral+cortex#imid_18309883
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000124334-IL9R/tissue/colon#imid_18299537
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000124334-IL9R/tissue/liver#imid_18299546
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000124334-IL9R/tissue/kidney#imid_18299531
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for IL9R IHC & IF
🧪Run testis first and expect round or early spermatids to stain (HPA: High in testis round or early spermatids); use adipose tissue as the negative tissue, with adipocytes at background (HPA: Not detected in adipocytes). On the testis slide, treat neighboring cells without visible signal as internal background comparators, but do not designate a particular cell type as IL9R-negative because the supplied HPA row identifies none (HPA: testis row).
Positive control tissue: Testis (Round or early spermatids, 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 IL9R; derive a cell-line control from the positive tissue's cell type (Round or early spermatids) and confirm it by RNA or western blot first.
Technical controls: Include a no-primary, secondary-only control; an isotype control matched to the primary antibody’s host species and clonality; and IL9R-knockout material if available, or competition with the antibody’s cognate immunogen peptide (standard IHC controls). For chromogenic testis sections, block endogenous peroxidase and, if using avidin–biotin detection, check for endogenous biotin (standard IHC practice).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected A07490-1 paraffin-section caption does not state a fixative (selected-SKU tissue-IHC caption). Retrieval dependence is unreported, so optimize antigen retrieval for the catalog antibody on paraffin sections (standard IHC practice). The supplied evidence does not establish whether frozen sections or IF/ICC are easier, and it documents no testis-specific artefact (HPA: no cell lines with ICC-IF images; supplied target/application evidence).
HPA tissue IHC evidence for IL9R
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Pending external verification. Presumed off target binding observed and disregarded.). 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 |
|---|---|---|---|---|
| Testis | Round or early spermatids | High | 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 → |
| Bone marrow | Hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Breast | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced IL9R IHC Tips
Troubleshoot IL9R staining in paraffin sections by checking retrieval, cell identity and compartment before interpreting signal intensity.
What should I change when IL9R staining is weak after retrieval?
Start with citrate pH 6.0 heat-induced retrieval at 95–98 °C for 20 min (page retrieval rule). Allow sections to cool in retrieval buffer, then compare a short dilution series against the reported 1:100 tissue-image condition without treating that image as evidence of a validated retrieval method (caption: A07490-1). If signal remains weak, test a different retrieval buffer or pH on adjacent sections while keeping detection and exposure conditions fixed (standard IHC practice). Judge each condition by signal in plausible IL9R-positive cells and background in a no-primary control, because stronger staining alone does not establish specificity (UniProt Q01113 localisation; standard IHC practice).
How can I assess whether fixation is masking IL9R in paraffin sections?
Target-specific IL9R sensitivity to fixation is unknown from the supplied evidence; the selected paraffin-section image reports no fixative (caption: A07490-1). Record the actual fixative, fixation duration and processing history for each specimen, then compare similarly handled sections before assigning weak signal to fixation (standard IHC practice). If matched material is available, vary retrieval while holding antibody dilution and chromogenic detection constant, and include a no-primary control in each run (standard IHC practice). Do not infer a preferred fixative from tissue staining patterns or receptor topology; those data describe expression and protein structure, respectively (HPA tissue IHC; UniProt Q01113 topology).
Where should convincing IL9R chromogenic staining appear?
Look first for cell-boundary staining, since IL9R has a transmembrane segment at residues 271–291 and is annotated at the cell membrane (UniProt Q01113 topology and localisation). Its extracellular region spans residues 41–270, while residues 292–521 are cytoplasmic, so the antibody epitope matters when evaluating accessibility after retrieval (UniProt Q01113 topology). A secreted localisation is also annotated, but diffuse extracellular chromogen alone cannot establish that the deposited material is IL9R (UniProt Q01113 localisation; standard IHC interpretation). Compare the staining pattern with cellular morphology and a no-primary section before calling broad cytoplasmic or extracellular colour positive (standard IHC practice).
Could epitope placement or IL9R isoforms explain discordant staining?
IL9R has 3 annotated isoforms, so check the antibody's stated immunogen against the sequence represented by each isoform before comparing sections or antibodies (UniProt Q01113 isoforms; standard IHC practice). The receptor contains an extracellular region at residues 41–270 and a cytoplasmic region at 292–521; accessibility can differ across these compartments in processed tissue (UniProt Q01113 topology; standard IHC practice). Glycosylation is annotated at residues 117 and 156, but the payload does not identify this antibody's epitope or establish an effect on staining (UniProt Q01113 glycosylation; caption: A07490-1). Compare epitope documentation and matched control sections before attributing discordance to splicing or glycosylation (standard IHC practice).
How should I follow up an IL9R IHC pattern with multiplex IF?
Use this as an orthogonal follow-up to chromogenic IHC, with an immune-cell marker chosen for the cells under study, because HPA describes IL9R expression in a subset of immune cells (HPA tissue IHC; standard IF practice). Choose fluorophores after checking the specimen's autofluorescence in unstained sections, and reserve a well-separated channel for a weak IL9R signal (standard IF practice). If the antibody recognises residues 292–521, assess permeabilisation for cytoplasmic access; an extracellular epitope within residues 41–270 may be accessible without it (UniProt Q01113 topology; standard IF practice). Confirm the epitope first, and compare single-stain and no-primary controls before interpreting apparent colocalisation (standard IF practice).
What controls help separate IL9R signal from chromogenic background?
Run a no-primary section and inspect pigment, damaged edges and extracellular deposits before adjusting the IL9R antibody concentration (standard IHC practice). Block endogenous peroxidase before an HRP/DAB detection step, and assess whether residual colour appears in the no-primary section (standard chromogenic IHC practice). Compare a dilution series around the reported 1:100 image condition on adjacent sections, judging cellular pattern as well as intensity (caption: A07490-1; standard IHC practice). HPA notes presumed off-target binding that was disregarded in its tissue assessment, so unexpected staining deserves independent antibody or orthogonal confirmation before it is called IL9R (HPA tissue IHC reliability description; standard IHC practice).
How should I score IL9R staining across heterogeneous tissue? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, then report the percentage of positive cells and intensity or an H-score from 0–300 using a consistent threshold (UniProt Q01113 localisation; standard IHC scoring practice). For scattered immune cells, positive-cell density per mm² can be more informative than a whole-section percentage (HPA tissue IHC profile; standard IHC scoring practice). Normalise to viable tissue area or to the number of eligible cells in the chosen compartment, and exclude necrosis and section folds using the same rules throughout (standard IHC scoring practice). Record retrieval, antibody dilution and detection batch alongside scores so technical changes are not mistaken for biological differences (standard IHC practice).
When should I doubt an apparent IL9R-positive cell?
Prioritise staining associated with plausible cells and membrane boundaries, given IL9R's membrane annotation and HPA's report of expression in a subset of immune cells (UniProt Q01113 localisation; HPA tissue IHC profile). HPA also reports high staining in round or early spermatids and low staining in lung macrophages, illustrating why the expected cell type depends on the sampled tissue (HPA tissue IHC). Treat staining confined to section edges, necrotic areas or a no-primary section as possible processing or detection artefact (standard IHC practice). Unexpected diffuse nuclear colour or widespread staining in cells reported as undetected warrants review of morphology, endogenous enzyme blocking and an independent specificity control before interpretation (UniProt Q01113 topology; HPA tissue IHC; standard IHC practice).
Boster reagents
Best IL9R / Interleukin-9 receptor IHC Antibodies
A07490-1 has a real IHC image from paraffin-embedded human liver carcinoma tissue (catalog image caption). Human, mouse, and rat reactivity is listed (catalog reactivity); no IF image is supplied (catalog images).
Real IHC data
Anti-IL-9R (L514) Antibody
Cat # A07490-1
A07490-1 is listed for IHC and WB in human, mouse, and rat (catalog applications and reactivity). Its IHC image shows paraffin-embedded human liver carcinoma tissue at 1:100 (catalog image caption).
Which to pick: Choose A07490-1 for paraffin-section tissue IHC: its image shows human liver carcinoma tissue at 1:100, and its listed IHC range is 1:50–1:200 (catalog image caption; catalog dilution). For cross-species IHC, A07490-1 lists mouse and rat reactivity, though its supplied IHC image is human only (catalog reactivity; catalog image caption). No IF/ICC application or image is supplied for A07490-1, so there is no supported IF/ICC pick here; the IHC caption does not report the fixative (catalog applications and images; catalog image caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q01113 (IL9R_HUMAN, Interleukin-9 receptor).
- Human Protein Atlas. IL9R tissue IHC expression (reliability: Approved).
- Human Protein Atlas. IL9R subcellular location (ICC-IF): Membrane.
- Human Protein Atlas. IL9R antibody validation summary (2 antibodies).
- Identification of Interleukin-9 Producing Immune Cells in Endometrial Carcinoma and Establishment of a Prognostic Nomogram. Frontiers in immunology 2020 — PMC7712424.
- STAT3/5-Dependent IL9 Overexpression Contributes to Neoplastic Cell Survival in Mycosis Fungoides. Clinical cancer research : an official journal of the American Association for Cancer Research 2016 — PMC4967550.
- PubMed PMID:1376929 — UniProt-cited evidence.
- PubMed PMID:8666384 — UniProt-cited evidence.
- PubMed PMID:10655549 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for IL9R (Interleukin-9 receptor). Expected localisation Cell membrane; secreted form also annotated (UniProt). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: A07490-1 at 1:50-1:200 (the product's own IHC validation image is shown). Controls (HPA): positive Testis (Round or early spermatids); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human liver carcinoma tissue; fixative not specified (datasheet A07490-1); 2. Fixation: Image fixative and duration unreported (datasheet A07490-1); 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-IL9R, 1:50-1:200 (datasheet A07490-1); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across sections (standard IHC practice; not target-specific). Selected-image fixative and duration unreported (datasheet A07490-1); verify before use.; Caveat: Presumed off-target binding may mimic IL9R staining (HPA tissue IHC); Isoform / epitope: 3 isoforms; check extracellular versus cytoplasmic epitope (UniProt); Regulation: Expression regulation unreported (UniProt). Evidence: UniProt Q01113, HPA tissue IHC · Approved, HPA ICC-IF · Membrane, Datasheet A07490-1, PMC4967550. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/IL9R/immunohistochemistry-ihc-guide.