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TLE2 / Transducin-like enhancer protein 2 · IHC design guide
Design Immunohistochemistry for TLE2
Plan TLE2 chromogenic IHC around the nuclear tissue pattern, with cerebellar Purkinje cells as a strong positive reference (HPA tissue IHC). This guide covers fixation, antibody dilution, detection and interpretation, including the reported off-target staining concern (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
TLE2 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 staining in tissues (HPA tissue IHC) | |
| Staining pattern | Nuclear staining across tissues, strongest in Purkinje cells (HPA tissue IHC) | |
| Antigen retrieval | Tris-EDTA pH 9.0 HIER, heat-mediated (datasheet A10582-1) | |
| Positive control | Cerebellum+4 more · see all | |
| Negative control | Adipose tissue+1 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep paraffin-section fixation consistent. (standard IHC practice; not target-specific) | |
| Caveat | Presumed off-target staining observed (HPA tissue IHC) | |
| Regulation | Tissue-dependent abundance (HPA tissue IHC) | |
| Isoform / epitope | 3 isoforms; verify epitope coverage for each (UniProt) |
Section 1
Recommended TLE2 IHC & IF Protocols
The catalog antibody protocol specifies heat retrieval in Tris-EDTA at pH 9.0 (datasheet A10582-1). The published IHC option uses paraffin-embedded xenograft sections (PMC5036407).
| Sample | Paraffin-embedded human tonsil tissue; fixative not specified (datasheet A10582-1) |
| Fixation | Image fixative and duration unreported (datasheet A10582-1); 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 A10582-1); 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-TLE2, 1:100-1:300 (datasheet A10582-1) |
| 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 | TLE2-positive staining in purkinje cells of cerebellum (HPA tissue IHC: High). HPA tissue profile: Nuclear expression in all tissues, most abundant in cerebellum. No signal in the no-primary control. |
💡Decision noteStart with Tris-EDTA pH 9.0 heat retrieval (datasheet A10582-1); the published xenograft protocol does not specify retrieval conditions (PMC5036407).
Section 2
What Is the Expected TLE2 Staining Pattern?
TLE2 is a nuclear transcriptional corepressor with no transmembrane segment (UniProt Q04725). In paraffin-section IHC, expect nuclear staining across tissues, strongest in cerebellar Purkinje cells (HPA: nuclear expression in all tissues; Purkinje cells High). Treat the pattern as a guide, not absolute specificity: HPA rates its tissue IHC reliability Enhanced but reports medium staining–RNA consistency and presumed off-target binding that was disregarded (HPA: reliability description).
| Strong, defined Purkinje-cell nuclei in cerebellum; some other nuclei stain less strongly. | This fits the most abundant reported tissue pattern (HPA: Purkinje cells High; nuclear expression in all tissues). Compare intensity within the same section before interpreting weaker neighboring cells as negative; HPA reports medium staining in several other specified cell types (HPA: tissue IHC). |
| The apparent signal is mainly cytoplasmic or membranous, with little nuclear staining. | Recheck localization and controls before calling this TLE2. A predominantly nonnuclear IHC pattern conflicts with the expected nucleus (UniProt Q04725: Nucleus; HPA: nuclear tissue expression). It could reflect background or off-target staining; appearance alone cannot identify its cause (general IHC interpretation). |
| Adipocytes or chondrocytes show convincing chromogenic staining. | These specific cell types were reported as Not detected (HPA: adipose-tissue adipocytes; soft-tissue chondrocytes). Review the negative control and whether the deposit lies in cells. Off-target binding or endogenous detection activity is possible (general IHC interpretation); the HPA result does not make every cell in those tissues negative. |
| Color spreads diffusely through tissue, obscuring cell boundaries and nuclei. | This is difficult to score as specific nuclear staining (HPA: nuclear tissue pattern). Diffuse color can arise from nonspecific detection or excess chromogen development (general chromogenic IHC practice). Compare a matched negative control and the nuclear counterstain before assigning TLE2 positivity. |
| Purkinje-cell nuclei show no detectable staining in an otherwise interpretable cerebellar section. | A missed expected positive warrants a run check (HPA: cerebellar Purkinje cells High). Inspect tissue preservation, detection and counterstain, then review the antibody’s documented IHC-P conditions (general IHC practice). Absence in one section cannot by itself establish biological TLE2 loss. |
💡Expected TLE2 appearanceCall a convincing positive when chromogen is concentrated in nuclei, especially strongly in cerebellar Purkinje cells; predominantly cytoplasmic color or staining in reported Not detected adipocytes or chondrocytes needs control review (HPA: tissue IHC; UniProt Q04725: Nucleus).
| Tissue and cell choice | Cerebellar Purkinje cells offer the strongest listed positive comparison (HPA: High). Respiratory epithelial cells in bronchus and neuronal cells in caudate or cortex are Medium (HPA: tissue IHC). A weaker result there should be judged against its own tissue context, not against Purkinje-cell intensity. |
| Compartment and protein architecture | TLE2 is nuclear and has no transmembrane segment or signal peptide (UniProt Q04725). The supplied record gives no basis to expect a membrane-bound or secreted-tissue pattern. Score defined nuclear staining first; a surface-like chromogenic outline requires investigation (general IHC interpretation). |
| Antibody evidence | HPA049103 has Enhanced IHC validation (HPA: antibody validation), while the tissue profile notes medium staining–RNA consistency and presumed off-target binding that was disregarded (HPA: reliability description). These qualifications support using cell type, compartment and controls together; validation does not make every colored structure specific. |
| Isoforms and modifications | UniProt lists three isoforms and four modified residues, including phosphorylated sites (UniProt Q04725). No epitope position or isoform-specific IHC behavior is supplied. Do not infer that one isoform, phosphorylation state or retrieval choice explains a staining difference from these annotations alone. |
| IF/ICC Q&A: should focal-adhesion signal be expected in paraffin IHC? | HPA reports mainly nuclear bodies and an additional focal-adhesion location in ICC-IF (HPA: subcellular ICC-IF). Its tissue IHC profile reports nuclear expression (HPA: tissue IHC). Treat focal-adhesion signal as an ICC-IF observation, not an established positive pattern for chromogenic paraffin IHC. |
| Situation | Likely cause | Next action |
|---|---|---|
| No nuclear signal in cerebellar Purkinje cells. | A positive expected to be High is missing (HPA: Purkinje cells High); the cause is undetermined from the slide alone. | Check a known-positive section and matched detection controls; review the catalog antibody’s IHC-P instructions and run records (general IHC practice). |
| Only cytoplasmic or membrane-like staining is visible. | The dominant compartment disagrees with the reported nuclear pattern (HPA: tissue IHC; UniProt Q04725: Nucleus). | Inspect nuclear counterstain and a negative control; reassess cell boundaries and background before scoring the slide (general IHC practice). |
| Adipocytes or chondrocytes stain strongly. | Those cell types are Not detected in the cited HPA tissues (HPA: tissue IHC); off-target or detection activity is possible (general IHC practice). | Confirm which cell contains the color, then compare a matched negative control; report the discordance rather than counting it as expected TLE2 staining. |
| Diffuse chromogen makes nuclear scoring unreliable. | Background or overdevelopment can hide a cellular pattern (general chromogenic IHC practice). | Compare negative controls and nuclear counterstain; review blocking, wash and chromogen-development steps under the assay’s established conditions (general IHC practice). |
| Color persists in a control lacking primary antibody. | Primary-independent detection activity or background may contribute (general chromogenic IHC practice). | Check the detection reagents and relevant endogenous-activity blocking for the detection system used; resolve control staining before interpreting TLE2. |
| A Medium HPA tissue appears faint beside a cerebellar positive. | Reported levels differ by cell type (HPA: Purkinje cells High; listed bronchial and neuronal cells Medium). | Compare the appropriate cell type and nuclear pattern within each tissue; check controls before calling a faint result technical failure (general IHC interpretation). |
Experiment evidence
IHC & IF images showing TLE2 staining
A10582-1 · IHCImmunohistochemical analysis of paraffin-embedded human tonsil. 1, Tris-EDTA, pH9.0 was used for antigen retrieval. 2 Antibody was diluted a
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000065717-TLE2/tissue/cerebral+cortex#imid_13994495
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000065717-TLE2/tissue/colon#imid_13994423
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000065717-TLE2/tissue/liver#imid_13994416
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000065717-TLE2/tissue/kidney#imid_13994417
HPA · ICC-IF MCF-7Human Protein Atlas immunofluorescence, MCF-7, antibody HPA049103; green = TLE2, blue = nucleus, red = microtubules.
HPA · ICC-IF MCF-7Human Protein Atlas immunofluorescence, MCF-7, antibody HPA049103; green = TLE2, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for TLE2 IHC & IF
🧪Run cerebellum first: Purkinje cells should stain (HPA: High in Purkinje cells). Run adipose tissue as the negative tissue, with adipocytes at background (HPA: Not detected in adipocytes); on the cerebellum slide, use cells lacking specific staining as internal background comparators without assuming a particular cell type is negative.
Positive control tissue: Cerebellum (Purkinje cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show TLE2 in MCF-7, RT-4, U2OS, with annotated localisation: Nuclear bodies (supported) (HPA subcellular).
Technical controls: Include a no-primary (secondary-only) control, a concentration-matched isotype control matched to the primary antibody’s host species and clonality, and a knockout or peptide-block control where available (standard IHC practice). For chromogenic cerebellum IHC, block endogenous peroxidase and check for tissue pigment before interpreting signal; for IF, assess tissue autofluorescence (standard IHC/IF practice).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected A10582-1 paraffin-section caption does not state a fixative (selected-SKU caption). The caption reports Tris-EDTA retrieval at pH 9.0 and primary antibody at 1:200 for its tonsil example; it does not establish that retrieval is required for cerebellum (selected-SKU caption). No supplied evidence establishes that frozen sections or IF are easier for TLE2; brain tissue autofluorescence can complicate IF interpretation (standard IF practice).
HPA tissue IHC evidence for TLE2
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Enhanced — Medium consistency between antibody staining and RNA expression data. 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 |
|---|---|---|---|---|
| Cerebellum | Purkinje cells | High | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | Medium | Protein (IHC) | HPA → |
| Caudate | Neuronal cells | Medium | Protein (IHC) | HPA → |
| Cerebral cortex | Neuronal cells | Medium | Protein (IHC) | HPA → |
| Colon | Endothelial cells | Medium | Protein (IHC) | HPA → |
Section 3
Advanced TLE2 IHC Tips
Troubleshoot chromogenic TLE2 IHC in paraffin sections using the catalog antibody’s tissue example and independent localisation evidence.
Which retrieval conditions should I try first for weak TLE2 staining in paraffin sections?
Start with heat-mediated retrieval in Tris-EDTA at pH 9.0 (datasheet A10582-1). The catalog antibody’s paraffin-section example used that buffer in human tonsil, followed by primary antibody at 1:200 overnight at 4 °C (caption A10582-1). If nuclear staining remains weak, check heating uniformity and section adhesion before comparing a different retrieval buffer as a fallback; keep antibody concentration and detection conditions constant during that comparison. Score nuclear staining separately from diffuse cytoplasmic colour because TLE2 is annotated as nuclear (UniProt Q04725) and tissue staining is predominantly nuclear (HPA tissue IHC).
How should I troubleshoot variable TLE2 staining when fixation histories differ?
Record the fixative, fixation duration and processing history for each block before comparing staining, because the selected paraffin-section caption does not state a fixative (caption A10582-1). Target-specific fixation sensitivity for TLE2 is unknown from the supplied evidence; nuclear localisation and phosphorylation annotations cannot establish it (UniProt Q04725). Run sections from differently processed blocks together with the same Tris-EDTA retrieval at pH 9.0 and the same detection batch (datasheet A10582-1). If intensity tracks processing history, report that association and assess morphology and internal nuclear staining before assigning a biological difference to TLE2.
What staining compartment is credible for TLE2 in tissue IHC?
Prioritise nuclear chromogen when identifying TLE2-positive cells: UniProt places TLE2 in the nucleus, and HPA reports nuclear tissue expression (UniProt Q04725; HPA tissue IHC). HPA cell imaging places it mainly in nuclear bodies and additionally at focal adhesion sites, but those observations come from subcellular imaging and need separate validation before explaining a tissue IHC pattern (HPA subcellular). Compare nuclei within the same section and examine whether colour follows cell borders, cytoplasm or extracellular material instead. Persistent staining outside nuclei warrants checks for background and antibody specificity, particularly because the HPA tissue assessment notes presumed off-target binding (HPA tissue IHC).
Can this IHC stain distinguish TLE2 isoforms or phosphorylation states?
Do not assign a chromogenic signal to a particular TLE2 isoform without an epitope map and isoform-specific validation; the record lists 3 isoforms (UniProt Q04725). TLE2 also has annotated phosphorylation sites, including CK2-associated serine 228 and CDK1-associated serine 249 and threonine 253 (UniProt Q04725). Those annotations alone do not show whether fixation or retrieval alters binding of this antibody. When staining changes after processing, compare matched sections with the catalog retrieval at pH 9.0, then seek independent specificity evidence before interpreting an isoform or phosphorylation change (datasheet A10582-1).
How can I compare TLE2 IHC with multiplex IF in the same tissue?
Use IF as a separately optimised comparison and multiplex TLE2 with a validated marker for the cell population of interest, such as Purkinje cells in cerebellum, where HPA reports high TLE2 staining (HPA tissue IHC). Choose a fluorophore channel with low background in the specimen, inspect unstained tissue for autofluorescence and use single-colour controls to check bleed-through. Permeabilise sufficiently for antibody access to the nuclear epitope; TLE2 has no annotated transmembrane segment and is nuclear (UniProt Q04725). Compare cell-level nuclear localisation across methods, while keeping the catalog’s pH 9.0 retrieval tied specifically to its paraffin-section IHC example (datasheet A10582-1).
How do I distinguish background colour from weak nuclear TLE2 staining?
Inspect a no-primary control, endogenous peroxidase blocking, wash quality and chromogen development when brown colour spreads beyond nuclei; these are general chromogenic IHC checks. The selected example used primary antibody at 1:200 overnight at 4 °C and secondary antibody at 1:200 for 45 minutes at room temperature (caption A10582-1). Shorten chromogen development or adjust blocking and washing one variable at a time if the control is coloured. Require a reproducible nuclear pattern before calling weak cells positive, since HPA reports nuclear staining and notes presumed off-target binding in its tissue assessment (HPA tissue IHC).
What should I score when comparing TLE2 IHC across specimens? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and anatomical region before scoring, then record the percentage of cells with nuclear chromogen and, if useful, an H-score using intensity categories 0–3 multiplied by their percentages; this is a standard IHC scoring convention. Normalise positive-cell counts to total evaluable cells of the same type, or report positive-cell density per mm² of viable tissue. Exclude folds, edges and necrosis consistently, and keep retrieval, detection and image thresholds identical across specimens. Report cell type alongside the score because HPA shows high staining in cerebellar Purkinje cells but no detected staining in adipocytes (HPA tissue IHC).
When should an apparent TLE2-positive IHC result be questioned?
Treat a reproducible nuclear signal in identifiable cells as the starting interpretation, consistent with UniProt nuclear localisation and HPA nuclear tissue expression (UniProt Q04725; HPA tissue IHC). Question strong cytoplasmic or extracellular colour, staining confined to section edges or necrotic regions, and colour in a no-primary control; review morphology and general endogenous-enzyme controls. A cell-type mismatch also merits review: HPA reports high staining in Purkinje cells and no detected staining in adipocytes or chondrocytes (HPA tissue IHC). Resolve unexpected patterns with an independent specificity check before making a biological claim, since HPA notes presumed off-target binding in its tissue assessment (HPA tissue IHC).
Boster reagents
Best TLE2 / Transducin-like enhancer protein 2 IHC Antibodies
A10582-1 has an IHC image from paraffin-embedded human tonsil (IHC image caption) and lists IF use and Human/Mouse reactivity (catalog: applications; reactivity). No IF image is provided (catalog: IF image alts).
Real IHC data
Anti-TLE2 Antibody
Cat # A10582-1
A10582-1 has an IHC image of paraffin-embedded human tonsil (IHC image caption). It lists IF as an application and Human/Mouse reactivity (catalog: applications; reactivity), but provides no IF image (catalog: IF image alts).
Which to pick: Choose A10582-1 for paraffin-section tissue IHC, supported by its human tonsil image; the fixative is unreported (IHC image caption). For IF, A10582-1 lists that application, but ICC validation and an IF image are unreported (catalog: applications; IF image alts). A10582-1 lists Human/Mouse reactivity and has a Rabbit host, but its clone is unreported and the IHC image shows human tissue only (catalog: reactivity; host; clone; IHC image caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q04725 (TLE2_HUMAN, Transducin-like enhancer protein 2).
- Human Protein Atlas. TLE2 tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. TLE2 subcellular location (ICC-IF): Mainly localized to the nuclear bodies. In addition localized to the focal adhesion sites..
- Human Protein Atlas. TLE2 antibody validation summary (1 antibodies).
- NDRG1 overexpression promotes the progression of esophageal squamous cell carcinoma through modulating Wnt signaling pathway. Cancer biology & therapy 2016 — PMC5036407.
- RNA binding protein DDX3X drives pancreatic cancer progression via the TLE2-MYL9 axis. Science advances 2025 — PMC12429019.
- Expression of Groucho/TLE proteins during pancreas development. BMC developmental biology 2008 — PMC2551604.
- PubMed PMID:1303260 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
- PubMed PMID:15057824 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for TLE2 (Transducin-like enhancer protein 2). Expected localisation Nuclear staining in tissues (HPA tissue IHC). Antigen retrieval: Tris-EDTA pH 9.0 HIER, heat-mediated (datasheet A10582-1). Validated IHC antibody: A10582-1 at 1:100-1:300 (the product's own IHC validation image is shown). Controls (HPA): positive Cerebellum (Purkinje cells); negative Adipose tissue. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Paraffin-embedded human tonsil tissue; fixative not specified (datasheet A10582-1); 2. Fixation: Image fixative and duration unreported (datasheet A10582-1); 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 A10582-1); 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-TLE2, 1:100-1:300 (datasheet A10582-1); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep paraffin-section fixation consistent. (standard IHC practice; not target-specific); Caveat: Presumed off-target staining observed (HPA tissue IHC); Isoform / epitope: 3 isoforms; verify epitope coverage for each (UniProt); Regulation: Tissue-dependent abundance (HPA tissue IHC). Evidence: UniProt Q04725, HPA tissue IHC · Enhanced, HPA ICC-IF · Mainly localized to the nuclear bodies. In addition localized to the…, Datasheet A10582-1, PMC5036407. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/TLE2/immunohistochemistry-ihc-guide.