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SYNE2 / Nesprin-2 · IHC design guide
Design Immunohistochemistry for SYNE2
Plan SYNE2 chromogenic IHC-P around nuclear-membranous staining in several cell types (HPA tissue IHC). Start the catalog antibody at 2–5 μg/ml (datasheet A02818-1) and interpret staining with isoform-dependent localization in mind (UniProt).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
SYNE2 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 membrane in several cell types (HPA tissue IHC) | |
| Staining pattern | Nuclear-membranous staining across several cell types (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A02818-1) | |
| Positive control | Colon+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 A02818-1) | |
| Caveat | Isoform-dependent localization may change the pattern (UniProt) | |
| Regulation | Expression varies by isoform (UniProt) | |
| Isoform / epitope | 13 isoforms; cytoplasmic and C-terminal epitopes may differ (UniProt) |
Section 1
Recommended SYNE2 IHC & IF Protocols
The catalog antibody’s paraffin IHC protocol (datasheet A02818-1) is accompanied by a published mouse embryo staining protocol (PMC6244730).
| Sample | Paraffin-embedded human penis squamous cell carcinoma tissue; fixative not specified (datasheet A02818-1) |
| Fixation | Image fixative and duration unreported (datasheet A02818-1); 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 A02818-1); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A02818-1) |
| Primary antibody | Rabbit anti-SYNE2, 2-5 μg/ml (datasheet A02818-1) |
| Primary incubation | Overnight at 4 °C (datasheet A02818-1) |
| Detection | HRP-conjugated secondary, DAB chromogen (datasheet A02818-1) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | SYNE2-positive staining in glandular cells of colon (HPA tissue IHC: High). HPA tissue profile: Nuclear membranous expression in several different cell types. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA retrieval at pH 8.0 (datasheet A02818-1). The published excerpt does not specify retrieval conditions (PMC6244730).
Section 2
What Is the Expected SYNE2 Staining Pattern?
SYNE2 should appear chiefly as a rim around nuclei in paraffin-section IHC: HPA reports nuclear membranous staining across several cell types (HPA tissue IHC: Enhanced; medium consistency with RNA data). Expect clear staining in colon glandular cells, kidney tubular cells, placental cytotrophoblasts, nasopharyngeal respiratory epithelium and basal skin cells (HPA tissue IHC: High). Its C-terminal membrane span anchors an isoform at the outer nuclear membrane, with most of that protein facing the cytoplasm (UniProt Q8WXH0 topology and subcellular location).
| A defined brown rim outlines nuclei in kidney tubules or colon glands (HPA tissue IHC: High in both cell types). | This matches the reported nuclear membranous pattern (HPA tissue IHC). Assess the rim in the specified cells, rather than scoring all nearby cells together; HPA's intensity calls apply to named cell types (HPA tissue IHC). |
| Cytoplasm stains strongly while nuclear outlines are indistinct (HPA tissue IHC: nuclear membranous profile). | This does not reproduce the principal tissue-IHC pattern (HPA tissue IHC). Cytoplasmic signal is biologically possible for shorter soluble isoforms, so compartment alone cannot prove artefact; compare morphology, antibody validation and controls (UniProt Q8WXH0 isoforms and subcellular location; standard IHC practice). |
| Strong staining appears mainly in adipocytes or bone-marrow hematopoietic cells (HPA tissue IHC: Not detected). | That distribution conflicts with the sampled HPA observations and warrants a specificity check, including detection controls (HPA tissue IHC; standard IHC practice). An HPA 'Not detected' call is an observation for those cells, rather than proof that every specimen must be negative (HPA tissue IHC). |
| A broad brown haze covers tissue and obscures cell borders or nuclear rims (standard IHC practice). | The section is difficult to interpret for SYNE2 localisation (HPA tissue IHC: nuclear membranous profile). Examine a no-primary control for detection background and review blocking, washing and chromogen development (standard IHC practice). |
| Kidney tubular cells and colon glandular cells show no visible rim under the same run (HPA tissue IHC: High). | These are useful positive reference cell populations, but their absence does not by itself identify the failure step (HPA tissue IHC; standard IHC practice). Check tissue preservation, antibody and detection performance, and the retrieval conditions used for that assay (standard IHC practice). |
💡Expected SYNE2 appearanceCall a convincing positive a distinct nuclear rim in the named HPA-high cells, particularly kidney tubules or colon glands; diffuse tissue-wide brown colour or predominant staining of HPA-undetected cell populations is suspect and needs controls (HPA tissue IHC; standard IHC practice).
| Membrane topology (UniProt Q8WXH0 topology). | The C-terminal segment spans residues 6835–6855; residues 1–6834 face the cytoplasm and 6856–6885 the perinuclear space (UniProt Q8WXH0 topology). A nuclear outline is consistent with this arrangement, but the record does not locate the antibody epitope (UniProt Q8WXH0 topology; supplied HPA antibody data). |
| Isoform distribution (UniProt Q8WXH0 subcellular location). | The record lists 13 isoforms and reports distinct compartments: membrane-bearing forms can occupy the nuclear envelope or other membranes, while shorter soluble forms occur in cytoplasm or nucleus (UniProt Q8WXH0 isoforms and subcellular location). Interpret secondary staining with the antibody's epitope coverage in mind (standard IHC practice). |
| Strength of tissue evidence (HPA tissue IHC; HPA antibodies). | HPA calls the tissue-IHC profile Enhanced but reports only medium consistency between antibody staining and RNA expression; both listed antibodies have Enhanced IHC status (HPA tissue IHC; HPA antibodies: HPA003435, HPA050204). Use the reported cell-level pattern as a reference, not an absolute cutoff for every specimen (HPA tissue IHC). |
| Retrieval and detection (standard IHC practice). | Optimize antigen retrieval and primary-antibody concentration against a positive reference section, then inspect a no-primary control for endogenous detection activity (standard IHC practice). The supplied HPA and UniProt records provide no SYNE2-specific retrieval condition or fixation-sensitivity result (supplied HPA and UniProt records). |
| IF/ICC Q&A: should the IHC rim appear in fluorescence images? (HPA subcellular ICC-IF). | HPA reports nuclear membrane localisation as uncertain, intermediate filaments as supported, and primary cilium localisation as uncertain in ICC-IF (HPA subcellular ICC-IF). Those cell-image observations can differ from the tissue-IHC profile; evaluate IF/ICC on its own guide page (HPA subcellular ICC-IF; HPA tissue IHC). |
| Situation | Likely cause | Next action |
|---|---|---|
| No stain in kidney tubular cells or colon glandular cells (HPA tissue IHC: High). | The assay may have missed a visible signal, or the selected section may not reproduce the reference observation (standard IHC practice; HPA tissue IHC). | Verify that the named cells are present; review antibody dilution, retrieval, incubation and detection with a positive reference section (standard IHC practice). No target-specific retrieval setting is supplied (supplied HPA and UniProt records). |
| Predominantly cytoplasmic stain with no clear nuclear rim (HPA tissue IHC: nuclear membranous profile). | Possible nonspecific staining or detection background; shorter soluble isoforms also occur in cytoplasm (standard IHC practice; UniProt Q8WXH0 subcellular location). | Compare with a no-primary control and the HPA cell-level pattern; assess any cytoplasmic signal separately rather than treating it as proof of the expected tissue-IHC result (standard IHC practice; HPA tissue IHC). |
| Brown signal in adipocytes or bone-marrow hematopoietic cells (HPA tissue IHC: Not detected). | The observed distribution differs from HPA's sampled pattern; nonspecific binding or endogenous detection activity is possible (HPA tissue IHC; standard IHC practice). | Check a no-primary control, blocking and detection chemistry; compare staining in a named HPA-high population on the same run (standard IHC practice; HPA tissue IHC). |
| Diffuse brown background obscures nuclear boundaries (standard IHC practice). | Excess primary or detection signal, inadequate washing, or endogenous activity can obscure compartment calls (standard IHC practice). | Review titration, washing and chromogen development; require a readable nuclear outline before scoring SYNE2 localisation (standard IHC practice; HPA tissue IHC: nuclear membranous profile). |
| Some cells show a rim, but intensity varies across tissue (HPA tissue IHC: several cell types). | HPA reports different levels by cell type, including High in kidney tubules and Medium in bronchial respiratory epithelium; its antibody–RNA consistency is medium (HPA tissue IHC). | Record cell identity and intensity separately. Compare like cells across sections and use the HPA levels as reference observations, not universal pass–fail thresholds (HPA tissue IHC; standard IHC practice). |
Experiment evidence
IHC & IF images showing SYNE2 staining
A02818-1 · IHCIHC analysis of Nesprin 2/SYNE2 using anti-Nesprin 2/SYNE2 antibody (A02818-1). Nesprin 2/SYNE2 was detected in a paraffin-embedded section
A02818-1 · IHCIHC analysis of Nesprin 2/SYNE2 using anti-Nesprin 2/SYNE2 antibody (A02818-1). Nesprin 2/SYNE2 was detected in a paraffin-embedded section
A02818-1 · IFIF analysis of Nesprin 2/SYNE2 using anti-Nesprin 2/SYNE2 antibody (A02818-1). Nesprin 2/SYNE2 was detected in an immunocytochemical section
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000054654-SYNE2/tissue/cerebral+cortex#imid_1238248
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000054654-SYNE2/tissue/colon#imid_1238166
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000054654-SYNE2/tissue/liver#imid_1238175
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000054654-SYNE2/tissue/kidney#imid_1238160
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA050204; green = SYNE2, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA050204; green = SYNE2, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for SYNE2 IHC & IF
🧪Run kidney first and require staining in tubular cells (HPA: High in kidney cells in tubules). Run adipose tissue as the negative comparator, focusing on adipocytes (HPA: Not detected in adipocytes); on the kidney slide, background-only staining in adjacent non-tubular cells can serve as an internal reference, but their SYNE2-negative status is unverified (HPA: kidney result specifies tubular cells).
Positive control tissue: Colon (Glandular cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show SYNE2 in A-431, MCF-7, U2OS, ASC52telo, RPTEC/TERT1, hTERT-RPE1 (serum starved), with annotated localisation: Nuclear membrane (uncertain), Intermediate filaments (supported) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only slide; a species- and class-matched rabbit IgG isotype control (selected caption: rabbit primary); and a matched SYNE2 knockout specimen or validated peptide-block control (standard IHC practice). For kidney sections, quench endogenous peroxidase before HRP/DAB detection and check endogenous biotin if using avidin–biotin detection (standard IHC practice; selected caption: HRP/DAB).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected A02818-1 paraffin-section caption does not state a fixative (selected caption: fixative unreported). The caption uses heat retrieval in EDTA at pH 8.0; whether SYNE2 staining depends on that condition is unreported (selected caption: EDTA heat retrieval). Neither frozen sections nor IF/ICC is shown to be easier in the supplied evidence; kidney tubular endogenous biotin can cause artefactual signal with avidin–biotin detection (standard IHC practice).
HPA tissue IHC evidence for SYNE2
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Enhanced — 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 |
|---|---|---|---|---|
| Colon | Glandular cells | High | Protein (IHC) | HPA → |
| Kidney | Cells in tubules | High | Protein (IHC) | HPA → |
| Nasopharynx | Respiratory epithelial cells | High | Protein (IHC) | HPA → |
| Placenta | Cytotrophoblasts | High | Protein (IHC) | HPA → |
| Skin | Cells in basal layer | 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 → |
| Caudate | Glial cells | Not detected | Protein (IHC) | HPA → |
| Cerebellum | Cells in granular layer | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced SYNE2 IHC Tips
Troubleshoot SYNE2 chromogenic IHC in paraffin sections by checking retrieval, staining compartment, controls and scoring; IF/ICC considerations appear in one entry.
How should I retrieve SYNE2 in paraffin sections when the nuclear rim is faint?
Use heat-mediated retrieval in EDTA at pH 8.0 first (datasheet A02818-1). The selected paraffin-section example used that buffer before 2 μg/ml primary antibody overnight at 4°C, so keep those conditions together for an initial comparison (datasheet A02818-1). Check whether nuclear-rim staining improves in a positive cell population, such as kidney tubule cells, while monitoring tissue integrity and diffuse background (HPA tissue IHC: High in kidney tubule cells; standard IHC practice). If staining remains weak, test an alternative retrieval buffer on adjacent sections, changing one condition at a time and retaining EDTA pH 8.0 as the reference (standard IHC practice).
Could fixation explain weak or uneven SYNE2 staining?
SYNE2-specific fixation sensitivity is unknown because the selected paraffin-section caption does not state a fixative (datasheet A02818-1). Compare sections with documented, consistent fixation and processing, and assess whether weak nuclear-rim staining tracks with those records rather than with cell type or section quality (standard IHC practice). Apply the same EDTA pH 8.0 retrieval and 2 μg/ml primary antibody conditions across that comparison (datasheet A02818-1). If staining varies within a section, inspect folds, detachment and poorly preserved areas before changing retrieval or antibody concentration; those features can confound chromogenic IHC interpretation (standard IHC practice).
Which SYNE2 staining patterns should count as expected localisation?
Prioritise a nuclear-membranous rim in suitable cells because that is the reported tissue-IHC pattern (HPA tissue IHC: nuclear membranous expression in several cell types). A C-terminal membrane-spanning region at residues 6835–6855 anchors relevant isoforms, while most of the outer nuclear membrane-associated protein faces the cytoplasm (UniProt Q8WXH0 topology and subcellular location). Cytoplasmic or intranuclear staining needs epitope and isoform context because shorter soluble isoforms can occupy those compartments (UniProt Q8WXH0 subcellular location). Compare the candidate pattern with surrounding cell morphology and a positive tissue control, recording rim, cytoplasmic and nucleoplasmic signals separately rather than combining them into one score (standard IHC practice).
How can epitope placement change the SYNE2 pattern I see?
Map the antibody immunogen or stated epitope against the 13 listed SYNE2 isoforms before assigning a staining pattern to a specific form (UniProt Q8WXH0 isoforms; standard IHC practice). Full-length SYNE2 has N-terminal calponin-homology domains at residues 31–136 and 181–286, plus a C-terminal KASH region at 6826–6885 (UniProt Q8WXH0 domains). Isoforms with the C-terminal transmembrane span can occupy organellar membranes, whereas shorter soluble isoforms may be cytoplasmic or nuclear (UniProt Q8WXH0 subcellular location). If the antibody epitope is undisclosed, report the observed compartment and antibody identity without claiming pan-isoform detection; use an independently validated epitope or orthogonal assay when isoform assignment matters (standard IHC practice).
How should I check a SYNE2 IHC pattern with multiplex IF?
Use IF/ICC as a separate validation experiment, multiplexing SYNE2 with a marker for the expected cell type and a nuclear counterstain to locate the rim (standard IF practice; HPA tissue IHC: nuclear membranous expression). Choose spectrally separated fluorophores after checking tissue autofluorescence and single-label controls, especially where a weak rim could be obscured (standard IF practice). Match permeabilisation to the documented antibody epitope: most of full-length SYNE2 lies on the cytoplasmic side, while residues 6856–6885 face the perinuclear space (UniProt Q8WXH0 topology). HPA reports intermediate-filament localisation as supported, nuclear-membrane localisation as uncertain, and primary-cilium localisation as uncertain in ICC/IF, so interpret each compartment with its own controls (HPA subcellular).
What should I change when SYNE2 DAB staining is diffuse?
Separate true compartmental staining from diffuse chromogen by examining a no-primary control and checking whether signal follows cell outlines or tissue edges (standard IHC practice). The selected example used 10% goat-serum blocking, 2 μg/ml rabbit primary antibody overnight at 4°C, a peroxidase-conjugated secondary for 30 minutes at 37°C, and DAB development (datasheet A02818-1). Include an endogenous-peroxidase block and control DAB development as general chromogenic workflow steps (standard IHC practice). If background persists, titrate the primary around the documented starting condition and compare adjacent sections, preserving any reproducible nuclear-rim signal while reducing diffuse deposition (standard IHC practice; HPA tissue IHC: nuclear membranous expression).
How should I score SYNE2 staining across cell populations? ⚠ ANSWER MARKED FOR VERIFICATION
Define the eligible cell population and nuclear-rim compartment before scoring, since tissue IHC reports nuclear-membranous SYNE2 in several cell types (HPA tissue IHC: nuclear membranous expression). Record the percentage of rim-positive eligible cells and, when intensity is reproducible, an H-score using 0–3 intensity categories; reserve positive-cell density per mm² for comparisons where sampled area and cell distribution are defined (standard IHC practice). Normalise percentages to all evaluable cells of the same type, and normalise density to evaluable tissue area rather than total section area (standard IHC practice). Keep cytoplasmic and nucleoplasmic signals separate, use matched acquisition and scoring rules, and exclude folds or necrosis from the denominator (standard IHC practice; UniProt Q8WXH0 subcellular location).
How can I distinguish a true SYNE2 signal from artefact?
Look for reproducible nuclear-rim staining in morphologically intact cells, then compare its distribution with reported high-staining populations such as colon glandular cells or kidney tubule cells (HPA tissue IHC: High in colon glandular cells and kidney tubule cells). Treat strong staining confined to tissue edges, necrotic regions or the no-primary control as suspect, and check endogenous peroxidase before attributing DAB deposit to SYNE2 (standard IHC practice). A purely diffuse pattern warrants epitope review because shorter soluble isoforms can occur in cytoplasm or nucleus, although tissue IHC chiefly reports a nuclear-membranous pattern (UniProt Q8WXH0 subcellular location; HPA tissue IHC: nuclear membranous expression). Interpret absence cautiously: HPA rates the tissue evidence “Enhanced” but reports only medium consistency between staining and RNA data (HPA tissue IHC: reliability description).
Boster reagents
Best SYNE2 / Nesprin-2 IHC Antibodies
Catalog antibody A02818-1 has IHC data from human paraffin sections and IF data from HeLa cells (catalog image captions); its listed reactivity is human and monkey (catalog: reactivity).
Real IHC data
Anti-Nesprin 2/SYNE2 Antibody ®
Cat # A02818-1
A02818-1 will render with its own IHC figure from a human penis squamous cell carcinoma paraffin section (catalog: IHC image caption). The same SKU lists IHC, IF and ICC applications and has IF data from HeLa cells (catalog: applications; IF image caption).
Which to pick: Choose A02818-1 for paraffin-section IHC: its own figure documents EDTA pH 8.0 retrieval and 2 μg/ml primary antibody in a human tissue section; the fixative is unreported (catalog: IHC image caption). For IF/ICC, A02818-1 is also listed for both applications, with an IF image from HeLa cells at 5 μg/ml (catalog: applications; IF image caption). For monkey samples, A02818-1 lists monkey reactivity, but its supplied IHC and IF images show human samples (catalog: reactivity; IHC and IF image captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q8WXH0 (SYNE2_HUMAN, Nesprin-2).
- Human Protein Atlas. SYNE2 tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. SYNE2 subcellular location (ICC-IF): Mainly localized to the nuclear membrane and intermediate filaments. In addition localized to the primary cilium..
- Human Protein Atlas. SYNE2 antibody validation summary (2 antibodies).
- Depletion of Nesprin-2 is associated with an embryonic lethal phenotype in mice. Nucleus (Austin, Tex.) 2018 — PMC6244730.
- PubMed PMID:12118075 — UniProt-cited evidence.
- PubMed PMID:11792814 — UniProt-cited evidence.
- PubMed PMID:12408964 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for SYNE2 (Nesprin-2). Expected localisation Nuclear membrane in several cell types (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A02818-1). Validated IHC antibody: A02818-1 at 2-5 μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Colon (Glandular cells); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human penis squamous cell carcinoma tissue; fixative not specified (datasheet A02818-1); 2. Fixation: Image fixative and duration unreported (datasheet A02818-1); 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 A02818-1); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A02818-1); 8. Primary antibody: Rabbit anti-SYNE2, 2-5 μg/ml (datasheet A02818-1); 9. Primary incubation: Overnight at 4 °C (datasheet A02818-1). Watch for: Fixation: Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A02818-1); Caveat: Isoform-dependent localization may change the pattern (UniProt); Isoform / epitope: 13 isoforms; cytoplasmic and C-terminal epitopes may differ (UniProt); Regulation: Expression varies by isoform (UniProt). Evidence: UniProt Q8WXH0, HPA tissue IHC · Enhanced, HPA ICC-IF · Mainly localized to the nuclear membrane and intermediate filaments.…, Datasheet A02818-1, PMC6244730. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/SYNE2/immunohistochemistry-ihc-guide.