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SIK3 / Serine/threonine-protein kinase SIK3 · IHC design guide
Design Immunohistochemistry for SIK3
Plan SIK3 chromogenic IHC on paraffin sections using the catalog antibody A30762 at 1:100–1:300 (datasheet). Compare nuclear tissue staining (HPA tissue IHC) with the cytoplasmic location annotated for SIK3 (UniProt).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
SIK3 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 in tissue (HPA tissue IHC); cytoplasmic (UniProt) | |
| Staining pattern | General nuclear staining across tissue cell types (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Adrenal gland+4 more · see all | |
| Negative control | None in HPA (detected in all 45 tissues); use no-primary + isotype controls |
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 | Tissue nuclear staining differs from the cytoplasmic annotation (HPA tissue IHC; UniProt) | |
| Regulation | Expression regulation unspecified (UniProt) | |
| Isoform / epitope | Four isoforms; epitope coverage is unspecified (UniProt) |
Section 1
Recommended SIK3 IHC & IF Protocols
Compare the catalog antibody’s IHC-P protocol with four published SIK3 IHC protocols (PMC6856590; PMC6825558; PMC4222365; PMC8411910).
| Sample | Paraffin-embedded human lung carcinoma tissue; fixative not specified (datasheet A30762) |
| Fixation | Image fixative and duration unreported (datasheet A30762); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat-induced epitope retrieval in citrate buffer, pH 6.0, 20 min at 95–98 °C (standard rule: cytoplasmic / membrane antigen) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% normal serum of the secondary host, 30 min, room temperature (standard) |
| Primary antibody | Rabbit anti-SIK3, 1:100 - 1:300 (datasheet A30762) |
| 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 | SIK3-positive staining in glandular cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: General nuclear expression. No signal in the no-primary control. |
💡Decision noteStart with citrate pH 6.0 retrieval at 95–98 °C for 20 min (page retrieval rule); published citrate conditions offer comparison points (PMC6856590; PMC8411910).
Section 2
What Is the Expected SIK3 Staining Pattern?
SIK3 is annotated in the cytoplasm, including puncta when bound to YWHAZ, and has no transmembrane segment (UniProt Q9Y2K2). In paraffin-section IHC, HPA instead reports general nuclear staining, with high staining in several glandular, hematopoietic, epithelial and neural cell populations (HPA tissue IHC). HPA rates the tissue profile Enhanced, while noting medium agreement with RNA data and pending external verification (HPA tissue IHC).
| Clear nuclear staining in adrenal or breast glandular cells, with restrained background. | This matches the reported general nuclear profile and high staining in those cells (HPA tissue IHC). Score the stained cell population as well as intensity; the HPA profile does not establish that every cell in a section must stain (HPA tissue IHC). |
| Predominantly cytoplasmic puncta, or staining confined to cell borders, with little nuclear signal. | Cytoplasmic puncta are biologically plausible through YWHAZ binding (UniProt Q9Y2K2), but they differ from the reported tissue IHC profile (HPA tissue IHC). Border-only staining is also unsupported by that profile; check morphology and controls before calling either pattern SIK3-positive. |
| Strong staining in cells outside the expected positive population, while adjacent expected cells are weak. | Compare cell identity with the tissue-specific HPA annotations before scoring (HPA tissue IHC). An unexpected pattern may reflect antibody cross-reactivity or endogenous detection activity (general IHC practice); morphology alone cannot distinguish those causes. |
| Uniform color across nuclei, stroma and spaces between cells. | A diffuse deposit obscures the cell-restricted nuclear pattern reported for SIK3 (HPA tissue IHC). In chromogenic IHC, incomplete blocking, excess detection reagent or inadequate washing can raise background (general IHC practice); interpret the slide only after controls clarify the signal. |
| No staining in a section containing a documented high-staining cell population. | Absence in, for example, adrenal glandular cells conflicts with the HPA high-staining reference (HPA tissue IHC). Review tissue identity, positive-control performance and the IHC workflow before concluding that the sample lacks detectable SIK3 (general IHC practice). |
💡Expected SIK3 appearanceCall a convincing positive when identifiable HPA high-staining cells show distinct nuclear chromogen with low background (HPA tissue IHC); widespread color without cell or compartment definition is suspect (general IHC practice).
| Choice of reference tissue and cell population | HPA reports high staining in adrenal, appendix and breast glandular cells; bone marrow hematopoietic cells; bronchial respiratory epithelial cells; caudate glial cells; cerebellar granular-layer cells; and cerebral-cortex endothelial cells (HPA tissue IHC). Adipocytes are listed as low (HPA tissue IHC). |
| Compartment evidence | General nuclear staining describes HPA tissue IHC, whereas UniProt places SIK3 in the cytoplasm and notes YWHAZ-associated puncta (HPA tissue IHC; UniProt Q9Y2K2). Record the observed compartment explicitly; neither source resolves this discrepancy for a particular section. |
| IHC antibody validation | HPA lists 2 rabbit polyclonal antibodies, HPA045245 and HPA048161, with Enhanced IHC status (HPA antibodies). The tissue profile still has medium RNA agreement and awaits external verification (HPA tissue IHC), so validation supports a comparison rather than certainty about each stained cell. |
| Protein form and epitope uncertainty | SIK3 has 4 isoforms, kinase and UBA domains, and several modified residues (UniProt Q9Y2K2). The supplied record gives no antibody epitope; these features cannot predict which isoforms stain or whether a particular modification changes IHC recognition. |
| Antigen retrieval and detection | Target-specific fixation effects are not established by the supplied assay evidence. Verify with a matched IHC source before attributing a result to fixation. |
| IF/ICC Q: Does its reported membrane location settle IHC scoring? | A: No. HPA's IF summary says membrane but provides no main location or cell-line images (HPA subcellular). Compare that limited summary with UniProt cytoplasmic localisation and the HPA nuclear tissue-IHC profile; interpret IF/ICC on its separate guide page (UniProt Q9Y2K2; HPA tissue IHC). |
| Situation | Likely cause | Next action |
|---|---|---|
| Nuclei stain in a reference tissue, but background prevents cell-level scoring. | Chromogen or detection background may obscure the HPA nuclear pattern (HPA tissue IHC; general IHC practice). | Compare reagent controls, then review blocking, washes and detection exposure; score only cells whose boundaries and nuclei remain interpretable (general IHC practice). |
| An HPA high-staining cell population is blank. | A failed staining run, unsuitable control section or missed target cell population can mimic a negative result (general IHC practice). | Confirm the named cells are present, inspect the run's positive control, and repeat with reviewed retrieval and detection settings before interpreting absence (HPA tissue IHC; general IHC practice). |
| Only cytoplasmic puncta are visible. | Puncta can fit the UniProt YWHAZ-associated localisation but differ from the HPA nuclear tissue profile (UniProt Q9Y2K2; HPA tissue IHC). | Document both compartment and cell type, compare a high-staining reference tissue, and avoid converting a plausible cellular location into an automatically validated IHC result (HPA tissue IHC). |
| Staining is strongest in an unexpected cell type. | The pattern may reflect cross-reactivity, endogenous activity or incorrect cell identification (general IHC practice). | Recheck morphology against the tissue's HPA cell annotation and inspect detection controls; retain uncertainty if the expected high-staining cells remain weak (HPA tissue IHC; general IHC practice). |
| Adipocytes stain as strongly as nearby high-staining reference cells. | HPA lists adipocytes as low, so this relative pattern deserves scrutiny rather than automatic acceptance (HPA tissue IHC). | Compare cells within the same section, examine background and controls, and report the departure from the HPA reference without treating low as an absolute negative control (HPA tissue IHC; general IHC practice). |
| A different antibody yields a different compartment or cell pattern. | The available records show Enhanced IHC status for 2 antibodies but do not guarantee identical staining in every specimen (HPA antibodies; HPA tissue IHC). | Compare matched sections, cell identities and controls; report each observed pattern and the nuclear-versus-cytoplasmic source disagreement instead of forcing agreement (HPA tissue IHC; UniProt Q9Y2K2; general IHC practice). |
Experiment evidence
IHC & IF images showing SIK3 staining
A30762 · IHCImmunohistochemistry analysis of paraffin-embedded human lung carcinoma tissue, using QSK Antibody. The picture on the right is blocked with
A05580-1 · IFImmunofluorescence of SIK3 in rat brain cells with SIK3 antibody at 20 μg/mL.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000160584-SIK3/tissue/cerebral+cortex#imid_15071014
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000160584-SIK3/tissue/colon#imid_15070948
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000160584-SIK3/tissue/liver#imid_15070941
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000160584-SIK3/tissue/kidney#imid_15070934
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for SIK3 IHC & IF
🧪Run bone marrow first; hematopoietic cells should stain strongly (HPA: High in hematopoietic cells). HPA lists no negative tissue and detects SIK3 in all 45 scored tissues, so use no-primary and isotype controls for the negative reference; any cells without chromogen on the positive slide should show counterstain alone, but they are not validated SIK3-negative cells (HPA: no negative tissue; standard IHC practice).
Positive control tissue: Adrenal gland (Glandular cells, HPA High)
Negative control tissue: None in HPA: SIK3 is detected in all 45 scored tissues. Use a no-primary (secondary-only) and an isotype control instead.
ICC-IF cell lines (HPA subcellular resource): HPA carries no ICC-IF cell line for SIK3; derive a cell-line control from the positive tissue's cell type (Glandular cells) and confirm it by RNA or western blot first.
Technical controls: Include a no-primary, secondary-only slide and a host-species-matched polyclonal control or subclass-matched monoclonal isotype control, according to the primary antibody's clonality (standard IHC practice). A SIK3 knockout specimen is a biological negative if available; the selected antibody also has a peptide-blocked tissue image (caption: A30762), and bone marrow sections need an endogenous peroxidase control when using peroxidase detection (standard IHC practice).
⚠️Feasibility: A target-specific fixation window and antigen-retrieval dependency are unreported in the supplied evidence; the selected paraffin-section caption does not state a fixative (caption: A30762). The evidence does not establish whether frozen sections or IF are easier for SIK3. In bone marrow, endogenous peroxidase can produce chromogenic background, so evaluate the no-primary slide alongside the stained section (standard IHC practice).
HPA tissue IHC evidence for SIK3
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Enhanced — Medium consistency between antibody staining and RNA expression data. Pending external verification.). 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 |
|---|---|---|---|---|
| Adrenal gland | Glandular cells | High | Protein (IHC) | HPA → |
| Appendix | Glandular cells | High | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | High | Protein (IHC) | HPA → |
| Breast | Glandular cells | High | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| None in HPA: SIK3 is detected in all 45 scored tissues. Use a no-primary (secondary-only) and an isotype control instead. | ||||
Section 3
Advanced SIK3 IHC Tips
Troubleshoot SIK3 staining in paraffin sections by checking retrieval, compartment, cell identity and controls before comparing signal across samples.
How should I retrieve SIK3 in paraffin sections when staining is weak?
Start with citrate buffer at pH 6.0, heated to 95–98 °C for 20 min (page retrieval setting). Let sections cool in buffer, then compare staining with an identically processed control section; cooling and matched processing help assess retrieval effects (standard IHC practice). If signal remains weak, test a longer heating time on adjacent sections while watching for tissue damage (standard IHC practice). SIK3 has no transmembrane segment and is reported in the cytoplasm, so assess staining in intact cells as well as overall intensity (UniProt Q9Y2K2 topology and subcellular annotation).
Could fixation explain inconsistent SIK3 staining between paraffin blocks?
Target-specific sensitivity of SIK3 to fixation is unknown from the supplied evidence; the catalog image identifies paraffin-embedded lung carcinoma but does not state its fixative (catalog antibody A30762 IHC caption). Record each block’s fixative, fixation duration and processing history before comparing staining (standard IHC practice). If those conditions differ, stain adjacent sections in one run using the same pH 6.0 citrate retrieval and detection steps (page retrieval setting; standard IHC practice). Compare cell morphology and staining distribution alongside intensity, since damaged or poorly preserved areas can mislead chromogenic interpretation (standard IHC practice).
Should I expect cytoplasmic or nuclear SIK3 staining?
UniProt places SIK3 in the cytoplasm and reports cytoplasmic puncta when it binds YWHAZ (UniProt Q9Y2K2 subcellular annotation). HPA tissue IHC instead describes general nuclear expression, while its separate subcellular summary says membrane, so the supplied localisation evidence conflicts (HPA tissue IHC profile; HPA subcellular summary). Score nuclear, diffuse cytoplasmic and punctate cytoplasmic signal separately on the same section rather than collapsing them into one intensity value (standard IHC practice). If a compartment dominates unexpectedly, inspect morphology and controls before assigning it to SIK3; compartment alone cannot resolve these conflicting annotations (standard IHC practice; cited localisation evidence).
Could SIK3 isoforms or phosphorylation alter what the antibody detects?
SIK3 has 4 annotated isoforms, but the supplied catalog caption does not identify the antibody epitope or establish which isoforms it recognises (UniProt Q9Y2K2 isoforms; catalog antibody A30762 IHC caption). Its kinase domain spans residues 66–317 and its UBA domain spans 344–384; several modified residues are also annotated (UniProt Q9Y2K2 domains and modified residues). Check the antibody’s documented immunogen before drawing isoform-specific conclusions from staining (standard IHC practice). If sections show different patterns, report those patterns as antibody staining unless epitope mapping and suitable controls support a narrower interpretation (standard IHC practice).
How can I compare SIK3 immunofluorescence with chromogenic IHC?
Treat IF/ICC as a separate assay: the supplied catalog image shows paraffin-section IHC and provides no IF/ICC fixation or permeabilisation conditions (catalog antibody A30762 IHC caption). For multiplex IF, pair SIK3 with a marker that identifies the cell population being assessed, and choose spectrally separated fluorophores after checking tissue autofluorescence (standard IF practice). Because SIK3 has no transmembrane segment and is annotated as cytoplasmic, evaluate permeabilisation when testing access to an intracellular epitope (UniProt Q9Y2K2 topology and subcellular annotation; standard IF practice). Compare compartments only after confirming that each assay’s controls and cell identification support the observed signal (standard IHC/IF practice).
What should I check when SIK3 DAB staining is widespread?
Run a no-primary control and inspect whether colour persists, particularly in blood-rich or damaged regions; residual endogenous peroxidase can produce DAB signal (standard chromogenic IHC practice). Apply a peroxidase block and use a matched blocking and detection workflow, then compare adjacent sections at the same DAB development time (standard chromogenic IHC practice). Because HPA describes low tissue specificity for SIK3 RNA, staining across several tissues is not, by itself, evidence of nonspecific antibody binding (HPA RNA specificity). Check whether background follows tissue edges, necrosis or folds, and whether the expected cellular staining remains distinguishable (standard IHC practice).
How should I quantify heterogeneous SIK3 staining in tissue sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, since cytoplasmic SIK3 and general nuclear tissue staining are both reported in the supplied sources (UniProt Q9Y2K2 subcellular annotation; HPA tissue IHC profile). Within a prespecified viable region, record the percentage of positive cells and intensity categories, then calculate an H-score if ordinal intensity is reproducible (standard IHC scoring practice). For spatial comparisons, report positive cells per mm² and normalise counts to the measured viable tissue area or relevant cell population (standard IHC quantification practice). Keep retrieval, imaging and scoring thresholds consistent across sections being compared (standard IHC practice).
How can I distinguish true SIK3 staining from an artefact?
Use the catalog antibody’s paraffin-section lung carcinoma image and its peptide-blocked companion as product-specific reference evidence, while noting that its fixative is unreported (catalog antibody A30762 IHC caption). Assess whether staining belongs to intact identified cells and record its compartment; UniProt reports cytoplasm, whereas HPA tissue IHC reports general nuclear expression (UniProt Q9Y2K2 subcellular annotation; HPA tissue IHC profile). Treat signal concentrated at cut edges, folds or necrosis cautiously, and compare it with a no-primary control for endogenous enzyme colour (standard chromogenic IHC practice). Report discordant compartments or cell populations explicitly rather than assigning every positive pixel to SIK3 (standard IHC interpretation practice).
Boster reagents
Best SIK3 / Serine/threonine-protein kinase SIK3 IHC Antibodies
Explore SIK3 antibodies with IHC data from paraffin-embedded human lung carcinoma tissue (A30762 image caption) and IF data from rat brain cells (A05580-1 image caption).
Real IHC data
Anti-QSK Antibody
Cat # A30762
Real IF data
Anti-SIK3 Antibody
Cat # A05580-1
A30762 is listed for IHC and IF in human and mouse samples, with an IHC image from paraffin-embedded human lung carcinoma tissue (A30762 applications, reactivity and image caption). A05580-1 is listed for IF in human, mouse and rat samples, with an IF image from rat brain cells (A05580-1 applications, reactivity and image caption).
Which to pick: Choose polyclonal A30762 for tissue IHC at 1:100–1:300; its image documents paraffin sections, but the fixative is unreported (A30762 dilution_raw and image caption). For ICC, choose A30762, which lists ICC and IF; for IF in rat brain cells, A05580-1 has image evidence at 20 μg/mL (A30762 applications; A05580-1 image caption). A05580-1 has the broader listed species reactivity—human, mouse and rat—while its supplied IF image documents rat cells only (A05580-1 reactivity and image caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q9Y2K2 (SIK3_HUMAN, Serine/threonine-protein kinase SIK3).
- Human Protein Atlas. SIK3 tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. SIK3 subcellular location (ICC-IF): Membrane.
- Human Protein Atlas. SIK3 antibody validation summary (2 antibodies).
- Downregulated Salt-inducible Kinase 3 Expression Promotes Chemoresistance in Serous Ovarian Cancer via the ATP-binding Cassette Protein ABCG2. Journal of Cancer 2019 — PMC6856590.
- An LKB1-SIK Axis Suppresses Lung Tumor Growth and Controls Differentiation. Cancer discovery 2019 — PMC6825558.
- Salt-inducible kinase 3, SIK3, is a new gene associated with hearing. Human molecular genetics 2014 — PMC4222365.
- Salt-Inducible Kinase 3 Promotes Vascular Smooth Muscle Cell Proliferation and Arterial Restenosis by Regulating AKT and PKA-CREB Signaling. Arteriosclerosis, thrombosis, and vascular biology 2021 — PMC8411910.
- PubMed PMID:14976552 — UniProt-cited evidence.
- PubMed PMID:16554811 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for SIK3 (Serine/threonine-protein kinase SIK3). Expected localisation Nuclear in tissue (HPA tissue IHC); cytoplasmic (UniProt). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: A30762 at 1:100 - 1:300 (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Glandular cells); negative None in HPA (detected in all 45 tissues); use no-primary + isotype controls. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human lung carcinoma tissue; fixative not specified (datasheet A30762); 2. Fixation: Image fixative and duration unreported (datasheet A30762); verify before use.; 3. Sectioning: 4–5 µm sections on charged slides (standard); 4. Deparaffinisation: Xylene, graded ethanol series to water (standard); 5. Antigen retrieval: Heat-induced epitope retrieval in citrate buffer, pH 6.0 , 20 min at 95–98 °C…; 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% normal serum of the secondary host, 30 min , room temperature (standard); 8. Primary antibody: Rabbit anti-SIK3, 1:100 - 1:300 (datasheet A30762); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across paraffin sections. (standard IHC practice; not target-specific); Caveat: Tissue nuclear staining differs from the cytoplasmic annotation (HPA tissue IHC; UniProt); Isoform / epitope: Four isoforms; epitope coverage is unspecified (UniProt); Regulation: Expression regulation unspecified (UniProt). Evidence: UniProt Q9Y2K2, HPA tissue IHC · Enhanced, HPA ICC-IF · Membrane, Datasheet A30762, PMC6856590, PMC6825558. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/SIK3/immunohistochemistry-ihc-guide.