SIRT2 / NAD-dependent protein deacetylase sirtuin-2 · IHC design guide

Design Immunohistochemistry for SIRT2

Plan chromogenic IHC on paraffin sections with the IHC-validated antibody PB9160 at 0.5–1 μg/mL (datasheet: IHC-P dilution). Assess cytoplasmic staining by cell type, using skeletal muscle as a positive reference (HPA tissue IHC).

Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use

Immunohistochemistry protocol sheet for SIRT2 (IHC for SIRT2): expected localisation Cytoplasmic tissue staining (HPA tissue IHC); mitotic chromatin association (UniProt), antibody PB9160, validated IHC image, and IHC protocol steps — Printable **SIRT2** IHC protocol sheet — expected localisation Cytoplasmic tissue staining (HPA tissue IHC); mitotic chromatin association (UniProt), antibody PB9160, controls and protocol steps. Open the full SIRT2 IHC guide →

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SIRT2 Immunohistochemistry Experimental Design Guide

Expected localisation, validated protocols, controls and antibodies — the at-a-glance facts below, then the full design guide.

Must know before staining

	Expected localisation	Cytoplasmic tissue staining (HPA tissue IHC); mitotic chromatin association (UniProt)
	Staining pattern	Cytoplasmic staining, most abundant in skeletal muscle (HPA tissue IHC)
	Antigen retrieval	EDTA pH 8.0 HIER, heat-mediated (datasheet PB9160)
	Positive control ⓘ	Smooth muscle+4 more · see all
	Negative control ⓘ	Adipose tissue+4 more · see all

Important caveats

Reasons your staining may differ from the expected pattern.

	Fixation	Keep formalin fixation consistent across sections. (standard IHC practice; not target-specific)
	Caveat	Antibodies may recognize different isoforms (HPA tissue IHC)
	Regulation	Reduced expression in many gliomas (UniProt)
	Isoform / epitope	Five isoforms; antibody epitope coverage may differ (UniProt; HPA tissue IHC)

Section 1

Recommended SIRT2 IHC & IF Protocols

The catalog antibody’s IHC-P protocol is accompanied by 3 published SIRT2 chromogenic IHC protocols (PMC12133431; PMC4411155; PMC12673322).

Recommended immunohistochemistry (IHC-P) protocol parameters
Sample	Paraffin-embedded human lung cancer tissue; fixative not specified (datasheet PB9160)
Fixation	Image fixative and duration unreported (datasheet PB9160); 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 PB9160); 20 min, 95–100 °C (standard)
Peroxidase block	3% H2O2, 10 min, room temperature (standard)
Blocking	10% goat serum (datasheet PB9160)
Primary antibody	Rabbit anti-SIRT2, 0.5-1μg/ml (datasheet PB9160)
Primary incubation	Overnight at 4 °C (datasheet PB9160)
Detection	Streptavidin-biotin complex (SABC), DAB chromogen (datasheet PB9160)
Counterstain	Hematoxylin, blue, dehydrate and mount (standard)
Expected result	SIRT2-positive staining in smooth muscle cells of smooth muscle (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in several tissues, most abundant in skeletal muscle. No signal in the no-primary control.

💡Decision noteStart with EDTA pH 8.0 heat retrieval for the catalog antibody (datasheet: PB9160); the published lung tissue protocol used citrate pH 6 with a different antibody (PMC4411155).

Section 2

What Is the Expected SIRT2 Staining Pattern?

In paraffin IHC, expect predominantly cytoplasmic SIRT2 staining, especially in skeletal muscle; smooth muscle cells and spermatogonia are reported as high, while several other cell types show medium staining (HPA tissue IHC). Nuclear or mitotic localization is biologically possible (UniProt Q8IXJ6 subcellular location). SIRT2 has no transmembrane segment (UniProt Q8IXJ6 topology). HPA rates tissue IHC as Approved but reports low consistency with RNA expression (HPA tissue IHC).

What am I looking at on my slide?
Clear cytoplasmic chromogen in smooth muscle cells or spermatogonia, with less signal in neighboring cells.	This fits the reported high staining in those cell types (HPA tissue IHC). Score the named cells separately from surrounding tissue; a whole-section average could hide the expected cell-specific pattern.
Cytoplasmic staining with some nuclear signal in a subset of cells.	Cytoplasm is the main tissue IHC pattern (HPA tissue IHC). SIRT2 can move to chromatin around G2/M and mitosis (UniProt Q8IXJ6 subcellular location). Assess nuclear staining against cell context and controls before treating it as unexpected.
Strong staining in adipocytes, marrow hematopoietic cells, or bronchial respiratory epithelium.	Those cell types are reported as not detected (HPA tissue IHC). Unexpected staining could reflect cross-reactivity or endogenous detection activity (general IHC practice); compare the signal with a no-primary control and the expected positive cells.
Broad, uniform chromogen across cells and extracellular areas, obscuring cell boundaries.	This does not resolve the reported cell-specific cytoplasmic pattern (HPA tissue IHC). Diffuse background can arise from nonspecific binding, insufficient washing, or endogenous detection activity (general IHC practice). Interpret intensity only after background is controlled.
No staining in smooth muscle cells or spermatogonia on an otherwise readable section.	Both are reported as high-staining cells (HPA tissue IHC). An absent signal warrants checking antibody performance and the IHC workflow before calling the sample SIRT2-negative (general IHC practice); HPA staining varies by antibody and may reflect different isoforms (HPA tissue IHC).

💡Expected SIRT2 appearanceCall a positive result when distinct, predominantly cytoplasmic chromogen appears in the expected cells, especially high-staining smooth muscle cells or spermatogonia; diffuse section-wide color or equally strong staining in reported not-detected cell types is suspect (HPA tissue IHC; general IHC practice).

How each factor affects the staining
Cell type and tissue context	HPA reports high staining in smooth muscle cells and spermatogonia, medium staining in cortical neurons and lung macrophages, and low staining in cardiomyocytes and kidney tubule cells (HPA tissue IHC). Compare like cell types when judging intensity.
Subcellular distribution	SIRT2 is usually cytoplasmic during the cell cycle but can associate with chromatin at G2/M and mitosis; centrosome, spindle, and midbody locations are also listed (UniProt Q8IXJ6 subcellular location). A nuclear signal needs cellular context.
Isoforms and antibody agreement	Five isoforms are listed (UniProt Q8IXJ6 isoforms). HPA cautions that CAB004573 and HPA011165 may target different isoforms; its tissue annotation uses CAB004573, and both antibodies are IHC Approved (HPA tissue IHC; HPA antibodies).
Strength of tissue evidence	HPA describes low consistency between antibody staining and RNA expression despite its Approved IHC rating (HPA tissue IHC). Treat tissue intensity as a guide for comparison, not a universal cutoff for every specimen or antibody.
IF/ICC Q: should SIRT2 be cytoplasmic only?	A: HPA reports supported cytosol and nucleolar localization, supported nucleoplasm as an additional location, and uncertain plasma-membrane localization (HPA subcellular ICC-IF). Interpret IF/ICC images within that assay's own guide; these locations do not redefine the paraffin IHC pattern.

Why is my staining missing, weak or wrong?
Situation	Likely cause	Next action
Expected positive cells are blank.	Antibody or detection performance may be inadequate (general IHC practice); the expected high-staining cells are smooth muscle cells and spermatogonia (HPA tissue IHC).	Check a positive control processed with the section, confirm the antibody's IHC-P validation and working conditions, and review detection reagents before assigning a biological negative (general IHC practice).
Signal is weak in heart muscle or kidney tubules.	Cardiomyocytes and kidney tubule cells are reported at low staining levels (HPA tissue IHC).	Use a reported high-staining cell type as the performance reference; score weak target-cell signal relative to local background and the positive control (HPA tissue IHC; general IHC practice).
Nearly every compartment has brown staining.	Diffuse signal may reflect nonspecific binding, inadequate washing, or endogenous enzyme activity with enzyme-based detection (general IHC practice).	Inspect a no-primary control, verify the appropriate blocking step for the detection system, and review wash and chromogen development conditions (general IHC practice).
Nuclear staining dominates while cytoplasm appears blank.	Nuclear SIRT2 can occur in particular cell-cycle contexts, but HPA describes cytoplasmic tissue expression overall (UniProt Q8IXJ6 subcellular location; HPA tissue IHC).	Compare with expected positive cells and controls, then check whether the nuclear signal follows recognizable cell morphology or instead tracks background (general IHC practice).
Reported not-detected cells stain as strongly as positive cells.	This conflicts with the cell-type pattern for adipocytes, marrow hematopoietic cells, or bronchial respiratory epithelium (HPA tissue IHC); cross-reactivity or endogenous detection activity is possible (general IHC practice).	Compare adjacent expected positive cells and a no-primary control; review antibody specificity and detection blocking before scoring those cells as positive (general IHC practice).
Two antibodies give different tissue patterns.	CAB004573 and HPA011165 may recognize different isoforms, and the HPA tissue annotation is based on CAB004573 (HPA tissue IHC).	Record which antibody produced each pattern and evaluate each against its own validation and controls; do not assume the HPA tissue annotation applies equally to both (HPA tissue IHC; general IHC practice).

Source: UniProt Q8IXJ6 Human Protein Atlas tissue Human Protein Atlas subcellular

Experiment evidence

IHC & IF images showing SIRT2 staining

IHC analysis of SIRT2 using anti-SIRT2 antibody (PB9160). SIRT2 was detected in a paraffin-embedded section of human lung cancer tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1 μg/ml rabbit anti-SIRT2 Antibody (PB9160) overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) (Catalog # SA1022) with DAB as the chromogen.

PB9160 · IHCIHC analysis of SIRT2 using anti-SIRT2 antibody (PB9160). SIRT2 was detected in a paraffin-embedded section of human lung cancer tissue. Hea

IHC analysis of SIRT2 using anti-SIRT2 antibody (PB9160). SIRT2 was detected in a paraffin-embedded section of rat brain tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1 μg/ml rabbit anti-SIRT2 Antibody (PB9160) overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) (Catalog # SA1022) with DAB as the chromogen.

PB9160 · IHCIHC analysis of SIRT2 using anti-SIRT2 antibody (PB9160). SIRT2 was detected in a paraffin-embedded section of rat brain tissue. Heat mediat

HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000068903-SIRT2/tissue/cerebral+cortex#imid_18755624

HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000068903-SIRT2/tissue/colon#imid_18755558

HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000068903-SIRT2/tissue/liver#imid_18755543

HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000068903-SIRT2/tissue/kidney#imid_18755552

HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA011165; green = SIRT2, blue = nucleus, red = microtubules.

← swipe / scroll for more images →

Real IHC / IF data behind the staining-pattern expectations in this section.

Sample controls for SIRT2 IHC & IF

🧪Run smooth muscle first and expect smooth muscle cells to stain (HPA: High in smooth muscle cells). Use adipose tissue as the negative comparator because adipocytes are not detected (HPA: Not detected in adipocytes); on the positive slide, morphologically distinct cells without specific staining provide an internal background reference, rather than a presumed SIRT2-negative cell type.

Positive control tissue: Smooth muscle (Smooth muscle cells, HPA High)

Negative control tissue: Adipose tissue (HPA Not detected)

ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show SIRT2 in A-431, U-251MG, U2OS, RPTEC/TERT1, hTERT-RPE1 (serum starved), with annotated localisation: Nucleoli (supported), Cytosol (supported) (HPA subcellular).

Technical controls: Include a no-primary (secondary-only) slide, a host-matched rabbit isotype control matched to the primary antibody’s clonality if known, and a SIRT2 knockout sample or a cognate-peptide block when available (caption: rabbit anti-SIRT2 antibody; standard IHC controls). Check endogenous peroxidase and biotin background in the positive tissue, especially because the illustrated detection uses a biotinylated secondary, streptavidin–biotin complex and DAB (selected PB9160 tissue-IHC caption; standard chromogenic IHC practice).

⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the fixative used for the selected paraffin-section image is unreported (selected PB9160 tissue-IHC caption). The illustrated IHC used heat-mediated retrieval in EDTA at pH 8.0, which supplies a starting condition but does not establish that retrieval is required for every specimen (selected PB9160 tissue-IHC caption). The supplied evidence does not establish whether frozen sections or IF are easier; assess chromogenic background from endogenous peroxidase and biotin in smooth muscle sections before scoring (selected PB9160 tissue-IHC caption; standard chromogenic IHC practice).

HPA tissue IHC evidence for SIRT2

Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Low consistency between antibody staining and RNA expression data. CAB004573 and HPA011165 may target different isoforms. Annotated protein expression is based on CAB004573.). 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
Smooth muscle	Smooth muscle cells	High	Protein (IHC)	HPA →
Testis	Spermatogonia cells	High	Protein (IHC)	HPA →
Adrenal gland	Glandular cells	Medium	Protein (IHC)	HPA →
Breast	Myoepithelial cells	Medium	Protein (IHC)	HPA →
Cerebral cortex	Neuronal cells	Medium	Protein (IHC)	HPA →

Undetected expression · recommended negative controls

Tissue	Cell type	Level	Evidence	Source
Adipose tissue	Adipocytes	Not detected	Protein (IHC)	HPA →
Appendix	Glandular cells	Not detected	Protein (IHC)	HPA →
Bone marrow	Hematopoietic cells	Not detected	Protein (IHC)	HPA →
Bronchus	Respiratory epithelial cells	Not detected	Protein (IHC)	HPA →
Caudate	Glial cells	Not detected	Protein (IHC)	HPA →

Section 3

Advanced SIRT2 IHC Tips

Troubleshoot SIRT2 staining in paraffin sections by checking retrieval, cellular distribution, controls, and scoring before interpreting differences between specimens.

Which retrieval conditions should I try first for weak SIRT2 staining?

Start with heat-mediated retrieval in EDTA pH 8.0 for the catalog antibody on paraffin sections (datasheet PB9160). The illustrated section used 1 μg/ml primary antibody overnight at 4°C, so first check that retrieval and antibody exposure match the demonstrated conditions (datasheet PB9160). Run matched sections through a controlled heating and cooling cycle, keeping section thickness and detection conditions constant (standard IHC practice). If staining remains weak, compare a cautiously adjusted heating duration on adjacent sections while monitoring tissue damage (standard IHC practice). Do not interpret stronger staining alone as improved specificity; compare the expected cytoplasmic pattern and control tissues (HPA tissue IHC).

Could fixation explain inconsistent SIRT2 staining between paraffin blocks?

Target-specific SIRT2 sensitivity to fixative type or duration is unknown from the supplied evidence; the illustrated paraffin-section caption does not report its fixative (datasheet PB9160). Record each block’s fixation history and compare sections processed with the same retrieval, antibody concentration, and detection run (standard IHC practice). If histories differ, assess morphology and staining in internal control cells before attributing an intensity change to SIRT2 expression (standard IHC practice). Avoid calling a fixation effect from tissue staining patterns or SIRT2 sequence features alone (HPA tissue IHC; UniProt Q8IXJ6). For new samples, standardize fixation and processing, then document those conditions alongside the staining result (standard IHC practice).

Is nuclear SIRT2 staining credible when most tissue staining appears cytoplasmic?

Cytoplasmic staining across several tissues is the reported tissue IHC pattern, with the strongest abundance in skeletal muscle (HPA tissue IHC). Nuclear signal can still be plausible: SIRT2 associates with chromatin around the G2/M transition and during mitosis (UniProt Q8IXJ6 subcellular). Score cytoplasmic and nuclear staining separately, and compare nuclear-positive cells with their morphology and cell-cycle context (standard IHC practice; UniProt Q8IXJ6 subcellular). A uniform nuclear pattern throughout a section deserves closer control review because it exceeds the usual tissue profile (HPA tissue IHC). Check adjacent sections and omission controls before treating unusual nuclear DAB as SIRT2 localization (standard IHC practice).

How can isoforms or epitope accessibility change the apparent tissue pattern?

SIRT2 has 5 annotated isoforms, and isoforms 1 and 2 differ in reported tissue expression (UniProt Q8IXJ6 isoforms; UniProt Q8IXJ6 tissue specificity). The supplied PB9160 caption does not identify its epitope or establish isoform selectivity, so an individual stained section cannot assign an isoform (datasheet PB9160). Compare the antibody’s documented epitope, if available, with isoform sequences before making such a claim (standard IHC practice). Retrieval can also change access to a fixed epitope, so evaluate adjacent sections under consistent detection conditions (standard IHC practice). HPA reports low staining–RNA consistency and possible isoform differences between its antibodies; treat cross-antibody disagreement cautiously (HPA tissue IHC).

How should I investigate SIRT2 localization with a complementary IF experiment?

For a complementary IF study, multiplex SIRT2 with a marker identifying the cell type under investigation; lung macrophages provide one reported SIRT2-positive population (HPA tissue IHC). Choose a spectrally separated, preferably far-red SIRT2 channel after checking the specimen’s autofluorescence in each channel (standard IF practice). SIRT2 has no transmembrane segment and has reported cytosolic and nuclear locations, so permeabilize to expose intracellular epitopes while preserving morphology (UniProt Q8IXJ6 topology; HPA subcellular). Include single-color and secondary-only controls to assess bleed-through and nonspecific fluorescence (standard IF practice). Compare compartment-resolved IF with chromogenic staining cautiously because the supplied PB9160 tissue example documents paraffin-section IHC, not IF performance (datasheet PB9160).

What should I check when SIRT2 DAB staining is diffuse?

First inspect a no-primary control and the section edges to separate detection background from cellular SIRT2 staining (standard IHC practice). The demonstrated PB9160 method uses a biotinylated secondary, streptavidin–biotin complex, and DAB, so inspect both secondary binding and endogenous biotin contributions (datasheet PB9160; standard IHC practice). Apply an appropriate peroxidase block and assess whether DAB appears in blood-rich or damaged areas without primary antibody (standard IHC practice). The illustrated section used 10% goat serum blocking and 1 μg/ml primary antibody; use these as documented reference conditions when optimizing background (datasheet PB9160). Judge improvement by preserved cell-specific cytoplasmic staining rather than lower overall color alone (HPA tissue IHC; standard IHC practice).

How should I quantify SIRT2 staining across heterogeneous sections? ⚠ ANSWER MARKED FOR VERIFICATION

Define the anatomical region and cell population before scoring, then exclude folds, necrosis, and section edges consistently (standard IHC practice). For cellular chromogenic signal, record the percentage at each intensity level and calculate an H-score from 0–300; report cytoplasmic and nuclear scores separately (standard IHC practice). Normalize percentage-positive estimates to viable cells of the chosen type, or express positive-cell counts per mm² of analyzable tissue (standard IHC practice). Include matched controls and identical imaging or scoring thresholds across runs (standard IHC practice). This separation matters because tissue IHC is mainly cytoplasmic, while cell-cycle-dependent chromatin localization is also reported (HPA tissue IHC; UniProt Q8IXJ6 subcellular).

When should a SIRT2-positive area be considered an artefact?

Check whether DAB follows intact cells and a plausible compartment: tissue IHC commonly shows cytoplasmic SIRT2, with cell-context-dependent nuclear localization reported (HPA tissue IHC; UniProt Q8IXJ6 subcellular). Compare cell identity with reported patterns, such as medium staining in lung macrophages and absent staining in bronchial respiratory epithelial cells (HPA tissue IHC). Edge-restricted color, necrotic debris, or signal persisting in a no-primary control supports a technical explanation (standard IHC practice). Review peroxidase blocking when pigment appears in blood-rich regions, and review the biotin-based detection controls used with PB9160 (datasheet PB9160; standard IHC practice). Confirm unusual patterns on a well-preserved adjacent section before assigning biological significance (standard IHC practice).

Boster reagents

Best SIRT2 / NAD-dependent protein deacetylase sirtuin-2 IHC Antibodies

The IHC-validated antibody has paraffin-section images from human lung cancer and rat and mouse brain (PB9160 IHC image captions); a separate antibody lists human and rat ICC/IF (M00433-1 catalog).

Real IHC data

Anti-SIRT2 Antibody ®

Cat # PB9160

View product →

PB9160 is the rendered card, with IHC images of paraffin sections from human lung cancer, rat brain and mouse brain (PB9160 IHC image captions). M00433-1 lists human and rat ICC/IF, but has no supplied IF image and does not render as a card here (M00433-1 catalog applications, reactivity and image alts).

Which to pick: Choose PB9160 for tissue IHC: its own captions document paraffin sections, while its catalog lists IHC reactivity with human, mouse and rat (PB9160 IHC image captions; PB9160 catalog applications and reactivity). Choose the rabbit monoclonal M00433-1 for human or rat IF/ICC based on its listed applications and reactivity; no IF image is supplied (M00433-1 catalog title, applications, reactivity and image alts). PB9160 is the cross-species IHC choice across all three listed species; its captions do not report the fixative (PB9160 catalog reactivity; PB9160 IHC image captions).

Each figure is that product's own IHC / IF validation image from its datasheet.

References

UniProt Consortium. UniProt entry Q8IXJ6 (SIR2_HUMAN, NAD-dependent protein deacetylase sirtuin-2).
Human Protein Atlas. SIRT2 tissue IHC expression (reliability: Approved).
Human Protein Atlas. SIRT2 subcellular location (ICC-IF): Mainly localized to the nucleoli and cytosol. In addition localized to the nucleoplasm and plasma membrane..
Human Protein Atlas. SIRT2 antibody validation summary (2 antibodies).
SIRT2 Regulates the SMARCB1 Loss-Driven Differentiation Block in ATRT. Molecular cancer research : MCR 2025 — PMC12133431.
Expression and clinical significance of SIRT2 in gastric cancer: a retrospective study. Annals of medicine and surgery (2012) 2025 — PMC12688857.
Expression of sirtuin 1 and 2 is associated with poor prognosis in non-small cell lung cancer patients. PloS one 2015 — PMC4411155.
Sirtuin 1 and Sirtuin 2 Gene Expressions in Colorectal Cancers. Journal of clinical practice and research 2025 — PMC12673322.
PubMed PMID:10381378 — UniProt-cited evidence.
PubMed PMID:10393250 — UniProt-cited evidence.
PubMed PMID:12065666 — UniProt-cited evidence.