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STK3 / Serine/threonine-protein kinase 3 · IHC design guide
Design Immunohistochemistry for STK3
Plan chromogenic STK3 IHC in paraffin sections around its general cytoplasmic tissue pattern (HPA tissue IHC). Interpret nuclear staining in light of caspase-dependent nuclear entry (UniProt).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
STK3 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 cytoplasmic tissue staining (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic staining in adipocytes and glandular cells (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A02224-2) | |
| Positive control | Adipose tissue+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 paraffin-section fixation consistent across samples (standard IHC practice; not target-specific) | |
| Caveat | Staining and RNA show medium consistency (HPA tissue IHC) | |
| Regulation | Caspase cleavage may increase nuclear signal (UniProt) | |
| Isoform / epitope | 2 isoforms; cleavage splits chains, so check the epitope (UniProt) |
Section 1
Recommended STK3 IHC & IF Protocols
The catalog antibody protocol uses EDTA pH 8.0 retrieval (datasheet: A02224-2). The published IHC protocols below report STK3 staining in canine osteosarcoma and human esophageal carcinoma (PMC12194233; PMC11322239).
| Sample | Paraffin-embedded human renal cancer tissue; fixative not specified (datasheet A02224-2) |
| Fixation | Image fixative and duration unreported (datasheet A02224-2); 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 A02224-2); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A02224-2) |
| Primary antibody | Rabbit anti-STK3, 2-5μg/ml (datasheet A02224-2) |
| Primary incubation | Overnight at 4 °C (datasheet A02224-2) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet A02224-2) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | STK3-positive staining in adipocytes of adipose tissue (HPA tissue IHC: Medium). HPA tissue profile: General cytoplasmic expression. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA pH 8.0 retrieval (datasheet: A02224-2); neither published IHC excerpt specifies a retrieval method (PMC12194233; PMC11322239).
Section 2
What Is the Expected STK3 Staining Pattern?
In paraffin-section IHC, expect a generally cytoplasmic STK3 pattern; HPA rates the tissue staining profile Approved, with medium consistency between antibody staining and RNA data (HPA: tissue IHC). Medium staining is reported in adipocytes, adrenal glandular cells, and bronchial respiratory epithelial cells (HPA: tissue IHC). Nuclear or centrosomal localization is biologically possible (UniProt Q13188: subcellular location). STK3 has no transmembrane segment (UniProt Q13188: topology).
| Cytoplasmic chromogen in adipocytes, adrenal glandular cells, or bronchial respiratory epithelial cells, with recognizable cellular boundaries. | This matches HPA's general cytoplasmic profile and its reported medium staining in those cells (HPA: tissue IHC). Compare cells within the same section as well as across sections; HPA reports low tissue specificity, so a difference between tissues alone does not establish antibody specificity (HPA: tissue IHC). |
| Predominantly membranous or extracellular staining, or nuclear staining without a credible cytoplasmic pattern. | A membrane or extracellular pattern is unexpected for STK3, which lacks a transmembrane segment (UniProt Q13188: topology). Nuclear staining needs context: UniProt reports nuclear localization and cycling of the caspase-cleaved form (UniProt Q13188: subcellular location). Check morphology, controls, and antibody specificity before calling any unusual compartment an artefact. |
| Strong staining in oral or vaginal squamous epithelium while the expected control cells stain weakly or inconsistently. | HPA reports low staining in those squamous epithelial cells (HPA: tissue IHC). A strong reversed pattern raises concern for cross-reactivity or endogenous detection activity, rather than proving either cause (general IHC practice). Inspect the detection-only control and repeat with an independently validated IHC antibody where available (general IHC practice). |
| Uniform haze, edge staining, or chromogen across tissue and empty areas, obscuring cell boundaries. | That distribution does not resemble HPA's cellular cytoplasmic profile (HPA: tissue IHC). Background can arise from nonspecific binding, inadequate washing, or detection chemistry (general IHC practice). Judge the section against a detection-only control before assigning positive cells, and avoid scoring diffuse haze as STK3 expression (general IHC practice). |
| No cellular signal in an adrenal gland or adipose section selected as a positive control. | HPA reports medium staining in adrenal glandular cells and adipocytes, so a blank control warrants a run-level check (HPA: tissue IHC). Confirm that the relevant cells are present and assess control performance, antibody conditions, retrieval, and detection (general IHC practice). A single blank section cannot establish biological absence. |
💡Expected STK3 appearanceCall a section positive when identifiable adipocytes or glandular or respiratory epithelial cells show chiefly cytoplasmic, approximately medium cellular staining (HPA: tissue IHC); diffuse haze or dominant membrane staining is suspect (HPA: tissue IHC; UniProt Q13188: topology).
| Compartment and processing | UniProt places STK3 in cytoplasm, nucleus, and centrosome and reports nuclear–cytoplasmic cycling of its caspase-cleaved form (UniProt Q13188: subcellular location). Its annotated full-length and cleaved chains differ (UniProt Q13188: processing). Thus, nuclear staining can be plausible, but these records do not predict how much of it an individual paraffin section should show. |
| Antibody epitope and isoforms | UniProt lists 2 isoforms and chains spanning residues 1–491, 1–322, and 323–491 (UniProt Q13188: isoforms; processing). An antibody's recognized epitope determines which forms it can detect (general immunoassay principle). The supplied records give no epitope for the IHC antibody, so a particular fragment or isoform cannot be assigned from staining. |
| Tissue and validation context | HPA calls the tissue IHC profile Approved while describing only medium consistency with RNA data; it lists medium staining in several cell types and low staining in others (HPA: tissue IHC). HPA marks HPA073281 and CAB025316 Approved for IHC, whereas HPA007120 is Approved for ICC (HPA: antibody validation). Keep antibody-specific validation tied to the assay used. |
| IF/ICC Q: should its pattern match tissue IHC? | A: HPA reports approved intermediate-filament and rods-and-rings localization in ICC-IF images from HaCaT and U2OS (HPA: subcellular ICC-IF). HPA separately describes general cytoplasmic staining in tissue IHC (HPA: tissue IHC). Treat the ICC-IF structures as an assay-specific observation, not a required chromogenic paraffin-section pattern. |
| Situation | Likely cause | Next action |
|---|---|---|
| Expected control cells are blank across the run. | An assay step may have failed; HPA reports medium staining in adrenal glandular cells and adipocytes (HPA: tissue IHC). | Verify control tissue identity and cellular preservation, then check antibody application, the chosen retrieval condition, and chromogenic detection against the assay's IHC instructions (general IHC practice). Reassess study sections only after the control produces interpretable cellular staining. |
| Signal is weak in every section, including the positive control. | Weak assay performance is plausible when the expected HPA medium-staining cells also lose signal (HPA: tissue IHC; general IHC practice). | Review antibody concentration, incubation, retrieval, detection reagents, and counterstain balance using the same positive control (general IHC practice). Adjust one condition at a time; the supplied records do not establish a STK3-specific retrieval or fixation response. |
| Brown signal appears in a detection-only control. | The signal can reflect endogenous detection activity or detection-reagent background rather than primary-antibody binding (general IHC practice). | Use detection-appropriate endogenous-activity controls or blocking and verify the chromogen step according to the detection system (general IHC practice). Reinterpret any matching tissue pattern only after that background is resolved. |
| Strong staining appears mainly at tissue edges or as a section-wide haze. | Edge artefact or nonspecific background can obscure the cellular cytoplasmic pattern HPA describes (HPA: tissue IHC; general IHC practice). | Inspect a detection-only control, washing, blocking, and reagent coverage; then score only well-preserved areas with clear cell boundaries (general IHC practice). Do not turn a diffuse section-wide signal into a positive-cell count. |
| Staining is mostly nuclear or appears as puncta. | UniProt permits nuclear and centrosomal STK3 localization and describes cycling of the caspase-cleaved form (UniProt Q13188: subcellular location). | Check whether puncta lie within identifiable cells and whether the positive control also shows credible cytoplasmic staining (HPA: tissue IHC; general IHC practice). Report the observed compartment; avoid attributing it to cleavage or phosphorylation without separate evidence. |
| Strong staining is concentrated in cells HPA rates low. | An unexpected pattern may reflect specimen biology, cross-reactivity, or detection background; HPA rates oral and vaginal squamous epithelial cells low, not absent (HPA: tissue IHC; general IHC practice). | Compare the cells with an HPA medium-staining control and a detection-only control (HPA: tissue IHC; general IHC practice). If the discrepancy persists, check antibody-specific IHC validation before interpreting it as changed STK3 expression. |
Experiment evidence
IHC & IF images showing STK3 staining
A02224-2 · IHCIHC analysis of STK3/MST-2 using anti STK3/MST-2 antibody (A02224-2). STK3/MST-2 was detected in paraffin-embedded section of human renal ca
A02224-2 · IFIF analysis of STK3/MST-2 using anti-STK3/MST-2 antibody (A02224-2). STK3/MST-2 was detected in immunocytochemical section of PC-3 cells. En
M02224 · IHCImmunohistochemical analysis of paraffin-embedded human kidney, using STK3 Antibody.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000104375-STK3/tissue/cerebral+cortex#imid_21247219
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000104375-STK3/tissue/colon#imid_21247161
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000104375-STK3/tissue/liver#imid_21247162
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000104375-STK3/tissue/kidney#imid_21247163
HPA · ICC-IF HaCaTHuman Protein Atlas immunofluorescence, HaCaT, antibody HPA007120; green = STK3, blue = nucleus, red = microtubules.
HPA · ICC-IF HaCaTHuman Protein Atlas immunofluorescence, HaCaT, antibody HPA007120; green = STK3, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for STK3 IHC & IF
🧪Run bronchus first and look for staining in respiratory epithelial cells (HPA: Medium in bronchus respiratory epithelial cells). HPA detects STK3 in all 45 scored tissues, so use no-primary and isotype controls for negatives; no cell population on the positive slide is established as STK3-negative, and cells lacking specific staining should show counterstain only (HPA: no negative tissue rows).
Positive control tissue: Adipose tissue (Adipocytes, HPA Medium)
Negative control tissue: None in HPA: STK3 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 ICC-IF images show STK3 in HaCaT, U2OS, with annotated localisation: Intermediate filaments (approved), Rods & Rings (approved) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only slide, a concentration-matched rabbit IgG isotype control, and, if available, an STK3 knockout biological control (caption: rabbit primary antibody). Quench endogenous peroxidase and check for endogenous biotin before using the caption’s biotin-based DAB detection system (caption: biotinylated secondary, SABC and DAB).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected A02224-2 paraffin-section caption does not state its fixative (caption: fixative not stated). The reported IHC conditions use heat retrieval in EDTA at pH 8.0; no supplied evidence establishes that frozen sections or IF are easier (caption: EDTA retrieval; HPA: ICC-IF images in HaCaT and U2OS). In bronchus, inspect mucus and section edges for nonspecific chromogen before scoring epithelial staining (HPA: bronchus respiratory epithelial cells).
HPA tissue IHC evidence for STK3
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — 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 |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Medium | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | Medium | Protein (IHC) | HPA → |
| Appendix | Glandular cells | Medium | Protein (IHC) | HPA → |
| Breast | Adipocytes | Medium | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | Medium | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| None in HPA: STK3 is detected in all 45 scored tissues. Use a no-primary (secondary-only) and an isotype control instead. | ||||
Section 3
Advanced STK3 IHC Tips
Troubleshoot STK3 staining in paraffin sections by checking retrieval, detection controls, cellular localisation and scoring before interpreting chromogenic signal.
What retrieval conditions should I try first for weak STK3 staining?
Start with heat-mediated antigen retrieval in EDTA at pH 8.0 for paraffin sections (datasheet A02224-2). The documented renal cancer section workflow then used 10% goat serum, 2 µg/ml primary antibody overnight at 4°C, and DAB detection (datasheet A02224-2). If staining remains weak, adjust retrieval heating or cooling in small steps while keeping section preparation and detection constant (standard IHC practice). Compare each condition with a matched no-primary control, because stronger retrieval can also increase nonspecific signal or damage tissue morphology (standard IHC practice). The caption reports paraffin embedding but does not state the fixative (datasheet A02224-2).
Could fixation explain inconsistent STK3 staining between sections?
Target-specific STK3 sensitivity to fixation is unknown from the supplied evidence; the selected tissue caption does not report a fixative (datasheet A02224-2). Record the fixative, fixation duration, section thickness, and time in storage for each specimen before comparing staining intensity (standard IHC practice). Process test sections in parallel with the documented EDTA pH 8.0 retrieval and 2 µg/ml primary antibody conditions (datasheet A02224-2). If signal varies, compare replicate sections with similar morphology and a common reference section in the same staining run (standard IHC practice). Treat any observed fixation effect as a finding from that comparison, rather than an established STK3 property.
Should STK3 staining appear in nuclei, cytoplasm, or cytoskeletal structures?
Expect a predominantly cytoplasmic tissue pattern when evaluating chromogenic sections (HPA: general cytoplasmic expression), while nuclear and centrosomal localisation are also reported (UniProt Q13188 subcellular location). A caspase-cleaved form cycles between nucleus and cytoplasm, and Thr-117 phosphorylation inhibits nuclear translocation (UniProt Q13188). HPA also reports intermediate filaments and rods and rings in cell imaging, which are distinct observations from its tissue IHC profile (HPA subcellular; HPA tissue IHC). Score nuclear and cytoplasmic staining separately in identifiable, intact cells (standard IHC practice). Check a no-primary section before assigning fine puncta or filament-like DAB deposits to STK3 (standard IHC practice).
How could STK3 processing or epitope position change my IHC result?
STK3 has 2 listed isoforms and can be cleaved into residues 1–322 and 323–491 (UniProt Q13188 processing; isoforms). Its kinase domain spans residues 27–278, while its SARAH domain spans 437–484 (UniProt Q13188 domains). The supplied caption does not identify the catalog antibody's epitope, so staining cannot be assigned to one isoform or cleavage product from these data (datasheet A02224-2). Check documented epitope mapping before making such an assignment, and compare staining in the same cellular compartment across sections (standard IHC practice). If testing another antibody, document its epitope and validate its IHC specificity independently (standard IHC practice).
How should I follow up a chromogenic STK3 pattern with IF?
Use a separate IF validation workflow, since the selected antibody evidence describes paraffin-section IHC with DAB detection (datasheet A02224-2). Multiplex STK3 with a validated marker for the cell population identified by morphology, and include single-label controls to check channel overlap (standard IF practice). Choose fluorophores after inspecting unstained tissue for autofluorescence, particularly in the intended detection channels (standard IF practice). STK3 has no transmembrane segment and is reported in cytoplasmic and nuclear compartments, so optimise permeabilisation for intracellular epitope access in fixed cells (UniProt Q13188 topology; subcellular location; standard IF practice). HPA reports IF images in HaCaT and U2OS cells, but those observations do not establish this catalog antibody's IF performance (HPA subcellular).
What should I adjust when STK3 DAB staining is diffuse or widespread?
First compare the tissue section with a no-primary control and inspect whether staining follows tissue edges, damaged areas, or endogenous pigment (standard IHC practice). The documented workflow used 10% goat serum blocking, 2 µg/ml primary antibody, a biotinylated secondary, a streptavidin–biotin complex, and DAB (datasheet A02224-2). Add an appropriate peroxidase block and assess endogenous biotin background when using this detection chemistry (standard IHC practice). If controls are clean but staining remains diffuse, titrate primary antibody and shorten DAB development while preserving the same EDTA pH 8.0 retrieval (standard IHC practice; datasheet A02224-2). Interpret widespread cytoplasmic staining cautiously because HPA reports general cytoplasmic STK3 expression (HPA tissue IHC).
How can I quantify STK3 staining across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, because HPA reports general cytoplasmic tissue staining and UniProt also lists nuclear localisation (HPA tissue IHC; UniProt Q13188). For intact cells, record the percentage staining at intensity 0–3 and calculate an H-score from 0–300, or report the percentage of positive cells (standard IHC practice). For spatial questions, count positive cells per mm² of viable, annotated tissue and report cytoplasmic and nuclear results separately (standard IHC practice). Normalise comparisons to the same cell population, viable tissue area, staining run, and reference section, using fixed thresholds for all samples (standard IHC practice). Record exclusions for folds, edges, and necrosis before scoring (standard IHC practice).
How do I distinguish credible STK3 signal from artefact?
Look for reproducible staining in intact cells, with cytoplasmic signal assessed against HPA's general tissue pattern and nuclear signal considered against UniProt localisation (HPA tissue IHC; UniProt Q13188). Review the cell type as well as intensity: HPA reports medium staining in adipocytes and several glandular cell populations, with low staining in listed squamous, stromal, and hematopoietic populations (HPA tissue IHC). Treat isolated staining at section edges, in necrotic areas, or outside identifiable cellular compartments as suspect until controls support it (standard IHC practice). A no-primary section helps identify endogenous enzyme or detection background, and matched reference sections help assess run-to-run consistency (standard IHC practice). HPA rates its tissue evidence Approved with medium staining–RNA consistency, so localisation alone cannot prove antibody specificity (HPA tissue IHC).
Boster reagents
Best STK3 / Serine/threonine-protein kinase 3 IHC Antibodies
Two anti-STK3 antibodies have human paraffin-section IHC images (A02224-2 and M02224 IHC captions); A02224-2 also has a PC-3 IF/ICC image (A02224-2 IF caption).
Real IHC data
Anti-STK3/MST-2 Antibody ®
Cat # A02224-2
Real IHC data
Anti-STK3/Mst2 Rabbit Monoclonal Antibody
Cat # M02224
A02224-2 will render with IHC in a human renal cancer paraffin section and has a separate IF/ICC image in PC-3 cells (A02224-2 image captions). M02224 will render with IHC in a human kidney paraffin section; IF/ICC is listed as an application, but no IF image is supplied (M02224 IHC caption; catalog applications and IF images).
Which to pick: For tissue IHC, A02224-2 offers a documented human renal cancer paraffin-section example with EDTA pH 8 retrieval and DAB detection; the fixative is unreported (A02224-2 IHC caption). For IF/ICC, choose A02224-2 for its PC-3 fluorescence example (A02224-2 IF caption). For listed mouse or rat reactivity, consider the rabbit monoclonal M02224, whose supplied IHC image shows human kidney paraffin tissue only and does not report the fixative (M02224 catalog reactivity and clone; M02224 IHC caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q13188 (STK3_HUMAN, Serine/threonine-protein kinase 3).
- Human Protein Atlas. STK3 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. STK3 subcellular location (ICC-IF): Localized to the rods & rings and intermediate filaments..
- Human Protein Atlas. STK3 antibody validation summary (3 antibodies).
- Comparison of Differentially Expressed Genes in Human and Canine Osteosarcoma. Life (Basel, Switzerland) 2025 — PMC12194233.
- STK3 kinase activation inhibits tumor proliferation through FOXO1-TP53INP1/P21 pathway in esophageal squamous cell carcinoma. Cellular oncology (Dordrecht, Netherlands) 2024 — PMC11322239.
- STK3 is a transcriptional target of YAP1 and a hub component in the crosstalk between Hippo and Wnt signaling pathways during gastric carcinogenesis. Molecular cancer 2025 — PMC12220525.
- PubMed PMID:8566796 — UniProt-cited evidence.
- PubMed PMID:8816758 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for STK3 (Serine/threonine-protein kinase 3). Expected localisation General cytoplasmic tissue staining (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A02224-2). Validated IHC antibody: A02224-2 at 2-5μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Adipose tissue (Adipocytes); 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 renal cancer tissue; fixative not specified (datasheet A02224-2); 2. Fixation: Image fixative and duration unreported (datasheet A02224-2); 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 A02224-2); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A02224-2); 8. Primary antibody: Rabbit anti-STK3, 2-5μg/ml (datasheet A02224-2); 9. Primary incubation: Overnight at 4 °C (datasheet A02224-2). Watch for: Fixation: Keep paraffin-section fixation consistent across samples (standard IHC practice; not target-specific); Caveat: Staining and RNA show medium consistency (HPA tissue IHC); Isoform / epitope: 2 isoforms; cleavage splits chains, so check the epitope (UniProt); Regulation: Caspase cleavage may increase nuclear signal (UniProt). Evidence: UniProt Q13188, HPA tissue IHC · Approved, HPA ICC-IF · rods & rings and intermediate filaments., Datasheet A02224-2, PMC12194233, PMC11322239. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/STK3/immunohistochemistry-ihc-guide.