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SMARCA5 / SWI/SNF-related matrix-associated actin-dependent regulator of chromatin subfamily A member 5 · IHC design guide
Design Immunohistochemistry for SMARCA5
Plan SMARCA5 chromogenic IHC on human paraffin sections using the catalog antibody at 2–5 μg/mL (datasheet A02687-1). Assess nuclear staining against tissue controls, such as colon glandular cells with high staining and adipocytes with no detected staining (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
SMARCA5 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 | General nuclear staining (HPA tissue IHC) | |
| Staining pattern | Nuclear staining across diverse tissue cell types (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A02687-1) | |
| Positive control | Appendix+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 fixation consistent across paraffin sections (standard IHC practice; not target-specific) | |
| Caveat | Adipocytes and cholangiocytes may show no staining (HPA tissue IHC) | |
| Regulation | Ubiquitously expressed (UniProt) | |
| Isoform / epitope | No isoforms annotated; no signal peptide or propeptide (UniProt) |
Section 1
Recommended SMARCA5 IHC & IF Protocols
The catalog antibody’s IHC-P protocol is presented alongside three published SMARCA5 IHC protocols with usable methods details (PMC12274346; PMC13378543; PMC11538284).
| Sample | Paraffin-embedded human appendix adenocarcinoma tissue; fixative not specified (datasheet A02687-1) |
| Fixation | Image fixative and duration unreported (datasheet A02687-1); 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 A02687-1); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A02687-1) |
| Primary antibody | Rabbit anti-SMARCA5, 2-5 μg/ml (datasheet A02687-1) |
| Primary incubation | Overnight at 4 °C (datasheet A02687-1) |
| Detection | HRP-conjugated secondary, DAB chromogen (datasheet A02687-1) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | SMARCA5-positive staining in glandular cells of appendix (HPA tissue IHC: High). HPA tissue profile: General nuclear expression. No signal in the no-primary control. |
💡Decision noteStart with the catalog antibody’s heat-mediated EDTA pH 8.0 retrieval (datasheet A02687-1); citrate pH 6.0 is a published alternative for testis sections (PMC13378543).
Section 2
What Is the Expected SMARCA5 Staining Pattern?
SMARCA5 is a nuclear chromatin-remodeling protein with no transmembrane segment (UniProt O60264). In paraffin-section IHC, expect predominantly nuclear staining across many cell types, including colon glandular cells and lymph-node germinal-center cells with high reported staining (HPA tissue IHC). HPA describes general nuclear expression, but rates its tissue IHC evidence “Supported,” with only medium consistency between antibody staining and RNA expression (HPA tissue IHC).
| Clear nuclear staining in colon glandular cells or lymph-node germinal-center cells, with recognizable tissue architecture. | This matches HPA’s high staining in those specific cell populations and its general nuclear profile (HPA tissue IHC). Score the relevant cells and their nuclei; a positive field alone does not show that every cell in the tissue should stain (HPA tissue IHC; general IHC practice). |
| Predominantly cytoplasmic, membranous, or extracellular color, with little nuclear signal. | That distribution conflicts with the nuclear and chromosome localization of SMARCA5 and its lack of a transmembrane segment (UniProt O60264). Treat it as suspect staining and check the detection controls and morphology before assigning a biological explanation (general IHC practice). |
| Strong staining in adipocytes or liver cholangiocytes, especially when the signal is outside nuclei. | HPA reports SMARCA5 as not detected in these named cell populations (HPA tissue IHC). Unexpected staining may reflect antibody cross-reactivity or endogenous detection activity; assess each possibility with controls (general IHC practice). HPA’s observation does not label the entire adipose or liver section negative (HPA tissue IHC). |
| A diffuse haze covers nuclei, cytoplasm, and surrounding tissue, obscuring cell boundaries. | The haze cannot establish the reported nuclear pattern (HPA tissue IHC; general IHC practice). Possible general IHC causes include excess primary or detection reagent, inadequate washing, and endogenous detection activity; interpret only after controls distinguish background from cell-associated signal (general IHC practice). |
| No nuclear signal in a colon glandular-cell or lymph-node germinal-center-cell positive-control region. | This disagrees with HPA’s high staining reports for those cells (HPA tissue IHC). Check that the expected cells are present, then review primary-antibody, retrieval, and detection steps as general IHC troubleshooting (general IHC practice). A single failed section cannot establish SMARCA5 absence (general IHC practice). |
💡Expected SMARCA5 appearanceCall a section positive when distinct nuclear chromogenic staining is visible in the relevant cells, with strong signal plausible in HPA high populations such as colon glandular cells; isolated cytoplasmic or diffuse tissue color is a suspect positive (HPA tissue IHC; UniProt O60264; general IHC practice).
| Cell population and scoring | Expression is broadly distributed but varies by cell type: HPA lists high staining in several glandular, squamous, hematopoietic, and germinal-center populations, low staining in breast and salivary glandular cells, and no detected staining in adipocytes or cholangiocytes (HPA tissue IHC). Compare like cells, not whole-organ averages (general IHC practice). |
| Evidence strength and antibody choice | HPA calls tissue IHC “Supported” and notes medium antibody-staining/RNA consistency (HPA tissue IHC). HPA008751 and CAB005227 each have “Supported” IHC status; HPA008751 also has “Supported” ICC status (HPA antibodies). These labels support a starting expectation, while controls remain necessary for a new assay (general IHC practice). |
| Nuclear distribution and cell state | UniProt places SMARCA5 in the nucleus and on mitotic chromosomes, and describes associations with replication foci during S phase and DNA-damage sites (UniProt O60264). Those annotations provide biological context; the supplied tissue IHC record supports a general nuclear pattern, not a required punctate pattern in paraffin sections (HPA tissue IHC). |
| IF/ICC Q&A: where should signal appear? | Mainly in the nucleoplasm, with additional localization to nucleoli fibrillar centers in HPA’s ICC-IF data (HPA subcellular ICC-IF). Use that localization to interpret an IF/ICC image; it is a separate application from the chromogenic paraffin-section IHC pattern described here (HPA subcellular ICC-IF; HPA tissue IHC). |
| Situation | Likely cause | Next action |
|---|---|---|
| Positive-control nuclei are blank. | The result conflicts with high staining reported for the chosen positive cell population (HPA tissue IHC); a missed or ineffective assay step is possible (general IHC practice). | Confirm the expected cells on the section, then review primary-antibody application, detection, and retrieval using the assay’s controls (general IHC practice). No SMARCA5-specific retrieval requirement is supplied by UniProt or HPA. |
| Color is chiefly cytoplasmic or membranous. | The compartment differs from SMARCA5’s nuclear localization (UniProt O60264); nonspecific staining or detection background is possible (general IHC practice). | Inspect a no-primary control, reassess the IHC-validated antibody’s staining pattern, and score only clearly localized nuclear signal (general IHC practice; HPA antibodies). |
| Adipocytes or cholangiocytes stain strongly. | HPA reports these particular cell types as not detected (HPA tissue IHC); cross-reactivity or endogenous detection activity may account for unexpected color (general IHC practice). | Compare the same cell type with a no-primary control and a high-staining positive-control population; keep cell identity and nuclear localization in the comparison (HPA tissue IHC; general IHC practice). |
| Background obscures nuclei throughout the section. | Excess reagent, insufficient washing, or endogenous activity can produce diffuse color in a chromogenic assay (general IHC practice). | Use a no-primary control to locate detection background, then review blocking, reagent concentration, and washes within the chosen assay workflow (general IHC practice). |
| Breast or salivary glandular cells show faint nuclear color. | HPA lists low staining in those cell populations, so a weak result can fit the reported tissue pattern (HPA tissue IHC). | Compare matched cell types and a high-staining control before judging the assay unsuccessful; preserve the distinction between faint nuclear signal and diffuse background (HPA tissue IHC; general IHC practice). |
| A dark field appears positive, but nuclei cannot be distinguished. | Overall color alone does not demonstrate the general nuclear expression reported by HPA (HPA tissue IHC; general IHC practice). | Assess tissue morphology and nuclear counterstain, compare the no-primary control, and record a positive interpretation only where signal can be assigned to nuclei in identifiable cells (general IHC practice). |
Experiment evidence
IHC & IF images showing SMARCA5 staining
A02687-1 · IHCIHC analysis of SNF2H/SMARCA5 using anti-SNF2H/SMARCA5 antibody (A02687-1). SNF2H/SMARCA5 was detected in a paraffin-embedded section of hum
A02687-1 · IHCIHC analysis of SNF2H/SMARCA5 using anti-SNF2H/SMARCA5 antibody (A02687-1). SNF2H/SMARCA5 was detected in a paraffin-embedded section of hum
A02687-1 · IFIF analysis of SNF2H/SMARCA5 and Tubulin beta using anti-SNF2H/SMARCA5 antibody (A02687-1) and anti-Tubulin beta antibody (M05613-4). SNF2H/
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000153147-SMARCA5/tissue/cerebral+cortex#imid_2917076
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000153147-SMARCA5/tissue/colon#imid_2916994
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000153147-SMARCA5/tissue/liver#imid_2916995
HPA · PancreasHuman Protein Atlas IHC, Pancreas. Source: https://www.proteinatlas.org/ENSG00000153147-SMARCA5/tissue/pancreas#imid_2916946
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA008751; green = SMARCA5, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA008751; green = SMARCA5, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for SMARCA5 IHC & IF
🧪Run appendix first and expect glandular cell nuclei to stain (HPA: High in appendix glandular cells; UniProt O60264: nuclear localization). Run adipose tissue as the negative comparator, where adipocytes are not detected (HPA: Not detected in adipocytes); no appendix cell type is documented as an internal negative, so unstained cells on that slide should be checked for background without assuming they lack SMARCA5 (HPA: appendix glandular cells High; UniProt O60264: ubiquitously expressed).
Positive control tissue: Appendix (Glandular cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show SMARCA5 in A-431, U-251MG, U2OS, KOLF2.1J, HEK293, with annotated localisation: Nucleoplasm (supported) (HPA subcellular).
Technical controls: Include a no-primary (secondary-only) slide, a host- and clonality-matched isotype control, and a SMARCA5 knockout control or validated peptide-block control (standard IHC practice; selected-SKU caption: rabbit primary antibody). For chromogenic appendix sections, block endogenous peroxidase and check for background before scoring nuclear DAB signal (standard IHC practice; selected-SKU caption: HRP/DAB detection).
⚠️Feasibility: A target-specific fixation window and fixation effect are unreported; the selected-SKU paraffin-section caption also leaves the fixative unreported (selected-SKU caption: fixative not stated). The caption uses heat retrieval in EDTA at pH 8.0, but does not establish that retrieval is required (selected-SKU caption: heat-mediated EDTA retrieval). ICC-IF images support an IF localization check in the listed cell lines, while relative ease versus frozen sections and appendix-specific artefacts are unreported (HPA subcellular: ICC-IF images and supported nucleoplasmic localization; selected-SKU caption: paraffin-section IHC).
HPA tissue IHC evidence for SMARCA5
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Supported — 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 |
|---|---|---|---|---|
| Appendix | Glandular cells | High | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | High | Protein (IHC) | HPA → |
| Cervix | Glandular cells | High | Protein (IHC) | HPA → |
| Colon | Glandular cells | High | Protein (IHC) | HPA → |
| Duodenum | Glandular cells | High | Protein (IHC) | HPA → |
Section 3
Advanced SMARCA5 IHC Tips
Troubleshoot chromogenic SMARCA5 IHC by checking nuclear staining, section processing and controls; use the IF guidance for multiplex imaging.
What retrieval should I try when SMARCA5 nuclear staining is weak?
Start with heat mediated retrieval in EDTA, pH 8.0, for paraffin sections (datasheet A02687-1). The illustrated section was incubated with 2 μg/mL primary antibody overnight at 4°C, so compare retrieval conditions before changing antibody concentration (caption A02687-1). If staining remains weak, vary heating time in small increments and process control and test sections together; excess heating can damage morphology (standard IHC practice). Judge improvement by crisp nuclear signal in glandular cells, which show high appendix staining, while checking that background does not rise (HPA: High in appendix glandular cells; HPA: general nuclear expression).
Can fixation account for weak SMARCA5 staining in paraffin sections?
The selected tissue caption does not state its fixative, so SMARCA5 specific fixation sensitivity is unknown (caption A02687-1: fixative not stated). Record fixative, fixation duration, section age and processing conditions for each specimen, then compare matched sections using the same retrieval and detection conditions (standard IHC practice). Delayed or uneven fixation can impair morphology and antigen detection generally, but it does not establish a particular SMARCA5 effect (standard IHC practice). Use an internal nuclear positive population and a separately processed positive control to distinguish a specimen processing problem from a staining run problem (HPA: general nuclear expression; standard IHC practice).
Which staining compartment should count as SMARCA5 positive?
Score predominantly nuclear staining: SMARCA5 is annotated in the nucleus and on chromosomes, and tissue staining has a general nuclear pattern (UniProt O60264: subcellular location; HPA: general nuclear expression). Nucleoplasmic signal is expected, with an additional nucleolar fibrillar center location reported by cell imaging (HPA: subcellular location). Mitotic chromosome staining can be plausible, but do not require replication foci or damage site patterns in routine paraffin sections (UniProt O60264: mitotic chromosomes, replication foci and damage site recruitment). Treat isolated diffuse cytoplasmic DAB as suspect; review morphology, counterstain and a primary antibody omission control before scoring it (standard IHC practice).
Could an isoform or altered epitope explain patchy nuclear staining?
The supplied record lists 0 annotated isoforms, so it does not support assigning patchy IHC staining to a named SMARCA5 isoform (UniProt O60264: isoforms). The antibody epitope is not specified here; its overlap with the ATP binding, helicase C terminal or SANT regions cannot be assumed (UniProt O60264: domains; caption A02687-1). Several modified residues are annotated, including phosphoserines at positions 66, 116 and 137, but their effects on this antibody are unknown (UniProt O60264: modified residues). Compare serial sections under matched retrieval and staining conditions before interpreting regional loss as an epitope or isoform effect (standard IHC practice).
How should I assess SMARCA5 in a multiplex IF experiment?
For a separate IF assay, pair SMARCA5 with an epithelial marker such as cytokeratin when assessing glandular cells, and include a nuclear counterstain (HPA: High in appendix glandular cells; standard IF practice). Select spectrally separated fluorophores and consider a far red channel for SMARCA5 if tissue autofluorescence obscures shorter wavelengths (standard IF practice). SMARCA5 is nuclear and has no transmembrane segment, so optimise permeabilisation to allow antibody access to its nuclear epitope after fixation (UniProt O60264: subcellular location and topology; standard IF practice). Confirm nuclear overlap and inspect single stain and no primary controls for bleed through and background before interpreting colocalisation (standard IF practice).
How can I reduce diffuse or misleading DAB background?
The selected paraffin section used 10% goat serum blocking, a peroxidase linked secondary antibody and DAB development (caption A02687-1). Add a peroxidase blocking step, then check wash stringency, secondary antibody background and DAB development time on matched sections (standard IHC practice). Compare a no primary control with the stained section to identify endogenous enzyme activity or detection reagent binding (standard IHC practice). Since the expected pattern is generally nuclear, diffuse cytoplasmic or stromal color should prompt review of section edges, damaged areas and reagent controls before calling it SMARCA5 (HPA: general nuclear expression; standard IHC practice).
How should I quantify SMARCA5 staining across specimens? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and nuclear scoring threshold before analysis, then report the percentage of positive nuclei or a nuclear H score (HPA: general nuclear expression; standard IHC practice). For an H score, multiply each intensity category by its percentage of cells and sum the products across the assessed population (standard IHC practice). Normalise positive counts to all evaluable nuclei of the same cell type, or report positive nuclei per mm² with the sampled area stated (standard IHC practice). Keep retrieval, exposure and DAB development consistent, and exclude necrotic or poorly preserved regions from both numerator and denominator (standard IHC practice).
When is an apparent SMARCA5 positive result likely to be artefactual?
A convincing result has nuclear signal in intact cells, consistent with SMARCA5 nuclear localisation and the reported general nuclear tissue pattern (UniProt O60264: subcellular location; HPA: general nuclear expression). Glandular cells in appendix can provide a useful positive reference, while adipocytes are reported as not detected and should not be treated as a universal negative control (HPA: High in appendix glandular cells; HPA: Not detected in adipocytes). Suspect edge limited staining, necrotic regions, diffuse cytoplasmic color or staining reproduced without primary antibody (standard IHC practice). Check a peroxidase block and no primary control before attributing residual DAB to SMARCA5 (standard IHC practice).
Boster reagents
Best SMARCA5 / SWI/SNF-related matrix-associated actin-dependent regulator of chromatin subfamily A member 5 IHC Antibodies
A02687-1 has human paraffin-section IHC images and an A549 IF image (catalog image captions); M02687 lists ICC/IF and Human, Mouse and Rat reactivity (catalog applications and reactivity).
Real IHC data
Anti-SNF2H/SMARCA5 Antibody ®
Cat # A02687-1
A02687-1 will render with an IHC image from human paraffin-embedded appendix adenocarcinoma; its catalog also shows IF in A549 cells (A02687-1 image captions). M02687 will render with listed ICC/IF applications and Human, Mouse and Rat reactivity, but no supplied IHC or IF image (M02687 catalog).
Which to pick: Choose A02687-1 for tissue IHC: its image documents staining of a human paraffin section at 2 μg/ml (A02687-1 IHC caption). For IF/ICC, A02687-1 has an A549 IF image at 5 μg/ml (A02687-1 IF caption); M02687 is a monoclonal option with listed ICC/IF use and Human, Mouse and Rat reactivity, though no IF image is supplied (M02687 catalog). The A02687-1 IHC caption supports paraffin sections; it does not report the fixative (A02687-1 IHC caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry O60264 (SMCA5_HUMAN, SWI/SNF-related matrix-associated actin-dependent regulator of chromatin subfamily A member 5).
- Human Protein Atlas. SMARCA5 tissue IHC expression (reliability: Supported).
- Human Protein Atlas. SMARCA5 subcellular location (ICC-IF): Mainly localized to the nucleoplasm. In addition localized to the nucleoli fibrillar center..
- Human Protein Atlas. SMARCA5 antibody validation summary (2 antibodies).
- SMARCA5 is required for the development of granule cell neuron precursors and Sonic Hedgehog Medulloblastoma growth. Scientific reports 2025 — PMC12274346.
- The role of micro RNAs let7c, 100 and 218 expression and their target RAS, C-MYC, BUB1, RB, SMARCA5, LAMB3 and Ki-67 in prostate cancer. Clinics (Sao Paulo, Brazil) 2013 — PMC3654318.
- PUM1 and PUM2 promote translation of chromatin regulators to ensure mammalian spermatogenesis. Science advances 2026 — PMC13378543.
- USP3 promotes DNA damage response and chemotherapy resistance through stabilizing and deubiquitinating SMARCA5 in prostate cancer. Cell death & disease 2024 — PMC11538284.
- PubMed PMID:9730600 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
- PubMed PMID:10880450 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for SMARCA5 (SWI/SNF-related matrix-associated actin-dependent regulator of chromatin subfamily A member 5). Expected localisation General nuclear staining (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A02687-1). Validated IHC antibody: A02687-1 at 2-5 μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Appendix (Glandular cells); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human appendix adenocarcinoma tissue; fixative not specified (datasheet A02687-1); 2. Fixation: Image fixative and duration unreported (datasheet A02687-1); 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 A02687-1); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A02687-1); 8. Primary antibody: Rabbit anti-SMARCA5, 2-5 μg/ml (datasheet A02687-1); 9. Primary incubation: Overnight at 4 °C (datasheet A02687-1). Watch for: Fixation: Keep fixation consistent across paraffin sections (standard IHC practice; not target-specific); Caveat: Adipocytes and cholangiocytes may show no staining (HPA tissue IHC); Isoform / epitope: No isoforms annotated; no signal peptide or propeptide (UniProt); Regulation: Ubiquitously expressed (UniProt). Evidence: UniProt O60264, HPA tissue IHC · Supported, HPA ICC-IF · Mainly localized to the nucleoplasm. In addition localized to the…, Datasheet A02687-1, PMC12274346, PMC13378543. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/SMARCA5/immunohistochemistry-ihc-guide.