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STK3 / Serine/threonine-protein kinase 3 · IHC design guide
Design Immunohistochemistry for STK3
Plan STK3 chromogenic IHC in paraffin sections using 2–5 μg/mL of the catalog antibody (datasheet A02224-2). Compare staining with the general cytoplasmic tissue pattern (HPA tissue IHC), while considering cleavage-related nuclear redistribution (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 across multiple tissue cell types (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 fixation consistent across paraffin sections. (standard IHC practice; not target-specific) | |
| Caveat | Caspase cleavage can shift STK3 into nuclei (UniProt) | |
| Regulation | Caspase cleavage favors nuclear signal (UniProt) | |
| Isoform / epitope | 2 isoforms; cleavage yields residues 1–322 and 323–491, so check epitope placement (UniProt) |
Section 1
Recommended STK3 IHC & IF Protocols
The catalog antibody’s IHC-P protocol is accompanied by published STK3 IHC methods for canine osteosarcoma (PMC12194233) and human esophageal squamous cell carcinoma (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 retrieval at pH 8.0 (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 chiefly cytoplasmic STK3 staining in the HPA-listed positive cell populations, including adipocytes, glandular cells and respiratory epithelial cells (HPA: general cytoplasmic expression; Medium staining). STK3 has no transmembrane segment, and UniProt also lists nuclear and centrosomal localization (UniProt Q13188 topology and subcellular location). HPA rates its tissue IHC profile Approved, with medium consistency between staining and RNA expression (HPA: tissue IHC reliability).
| Cytoplasmic staining in adipocytes, glandular cells or respiratory epithelial cells. | This matches HPA's general cytoplasmic profile and its reported Medium staining in these cell types (HPA: tissue IHC). Judge the stained cells against the anatomy on the same section; HPA's Approved rating carries a medium-consistency caveat for agreement with RNA expression (HPA: tissue IHC reliability). |
| Predominantly nuclear staining, with little cytoplasmic signal. | Check the pattern before calling it STK3: HPA describes general cytoplasmic tissue staining (HPA: tissue IHC). Nuclear STK3 is possible because the caspase-cleaved form cycles between nucleus and cytoplasm (UniProt Q13188 subcellular location); nuclear staining alone cannot establish cleavage or exclude artefact. |
| Strong staining in a cell population expected to stain weakly. | HPA reports Low staining in oral and vaginal squamous epithelial cells, ovarian stroma, smooth muscle, germinal-center cells and bone-marrow hematopoietic cells (HPA: tissue IHC). A strong result there merits checks for antibody cross-reactivity or endogenous detection activity (general IHC practice), rather than automatic assignment to STK3. |
| Diffuse color across cells, extracellular spaces and the section background. | The distribution does not resemble HPA's general cytoplasmic expression profile (HPA: tissue IHC). In chromogenic IHC, diffuse color may arise from nonspecific binding or detection background (general IHC practice); inspect a matched no-primary control before scoring cellular STK3 staining. |
| No detectable staining in an HPA-listed Medium-staining population. | Check tissue identity, antibody identity and the staining run before interpreting absence: HPA reports Medium staining in adipocytes, glandular cells and respiratory epithelial cells at specified sites (HPA: tissue IHC). HPA provides no negative tissue in this payload, so a blank section alone does not establish biological absence. |
💡Expected STK3 appearanceA credible positive is chiefly cytoplasmic, approximately Medium-intensity staining in an HPA-listed positive cell population; indiscriminate color outside those cells or across extracellular background is suspect (HPA: tissue IHC; general IHC practice).
| Tissue and cell selection | HPA reports Medium staining in specified cells of adipose tissue, adrenal gland, appendix, breast, bronchus and brain regions, and Low staining in several other listed cell populations (HPA: tissue IHC). Use the named cell type within each tissue when comparing slides; the tissue name alone is insufficient. |
| Evidence and antibody scope | The tissue profile is Approved with medium staining–RNA consistency (HPA: tissue IHC reliability). HPA073281 and CAB025316 are Approved for IHC, while HPA007120 has an ICC approval and no IHC status in this payload (HPA: antibodies). Those designations should remain application-specific. |
| Compartment and processing | UniProt places STK3 in cytoplasm and nucleus, and at the centrosome; its caspase-cleaved form cycles between nucleus and cytoplasm (UniProt Q13188 subcellular location and processing). Interpret an unusual nuclear pattern with that qualification, without claiming that an IHC image identifies a cleavage product. |
| Isoforms and antibody epitope | UniProt lists two isoforms and full-length and processed chains, including residues 1–322 and 323–491 (UniProt Q13188 isoforms and processing). The supplied record gives no epitope for the catalog antibody, so these entries cannot predict which forms it detects or justify a specific retrieval condition. |
| IF/ICC question: should its pattern match tissue IHC? | HPA reports approved ICC-IF localization to intermediate filaments and rods & rings, with images from HaCaT and U2OS (HPA: subcellular ICC-IF). Tissue IHC instead has a general cytoplasmic profile (HPA: tissue IHC). Treat the IF finding as application-specific localization evidence, not an IHC scoring requirement. |
| Situation | Likely cause | Next action |
|---|---|---|
| An HPA-listed Medium-staining cell population is blank. | The run, tissue identification or antibody application may need checking; HPA reports Medium staining for specific cell populations, not every cell in their tissues (HPA: tissue IHC). | Verify the annotated cell population and the antibody's IHC status, then review the run's controls and detection steps (HPA: antibodies; general IHC practice). Do not infer an STK3-specific fixation or retrieval effect from this result. |
| The section shows nuclear-only staining. | HPA's tissue profile is generally cytoplasmic, although UniProt permits nuclear localization of a caspase-cleaved form (HPA: tissue IHC; UniProt Q13188 subcellular location). | Compare the cytoplasmic signal and matched controls, and record nuclear staining separately (general IHC practice). Do not score the nuclear pattern as cleavage without independent evidence. |
| Strong color appears in an HPA-listed Low-staining cell population. | Cross-reactivity or endogenous detection activity could contribute (general IHC practice); HPA assigns Low staining to the specified populations (HPA: tissue IHC). | Confirm cell identity and inspect no-primary and detection controls (general IHC practice). Compare only like cell types; do not treat the HPA Low category as proof that all staining is false. |
| Color covers the whole section or extracellular areas. | Nonspecific binding or chromogenic detection background can obscure a cellular pattern (general IHC practice). | Inspect a no-primary control and review blocking, antibody dilution and detection conditions (general IHC practice). Score STK3 only where a cellular pattern can be distinguished from background. |
| Different tissues show different staining strengths. | HPA records both Medium and Low cell-specific staining and rates staining–RNA consistency as medium (HPA: tissue IHC). UniProt describes high expression in adult kidney, skeletal muscle and placenta, a separate tissue-specificity statement (UniProt Q13188). | Document tissue, cell type and compartment for each comparison (general IHC practice). Keep HPA's observed IHC levels distinct from UniProt's expression statement when interpreting differences. |
| IF/ICC images show filaments or rods and rings unlike the IHC section. | HPA assigns those structures to approved ICC-IF localization, while describing tissue IHC as generally cytoplasmic (HPA: subcellular ICC-IF and tissue IHC). | Use the tissue IHC profile to interpret the paraffin section and report the IF/ICC observation separately (HPA: tissue IHC and subcellular ICC-IF). |
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 score respiratory epithelial cells, which have Medium STK3 staining (HPA: bronchus, respiratory epithelial cells, Medium). HPA detects STK3 in all 45 scored tissues, so there is no HPA-supported negative tissue; use no-primary and isotype controls, and treat cells without visible staining on the positive slide as background references rather than validated STK3-negative cells (HPA: no negative rows; STK3 detected in all 45 scored tissues).
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 section; a rabbit IgG control matched to the primary antibody’s class and concentration; and STK3-knockout material as a biological negative (caption: rabbit primary antibody; standard IHC practice). Block endogenous peroxidase for DAB detection and assess background from bronchial mucus or inflammatory cells on the control sections (caption: DAB detection; standard IHC practice).
⚠️Feasibility: No target-specific fixation window or fixation effect is reported, and the selected A02224-2 paraffin-section caption does not state the fixative (caption: fixative not stated). That caption reports heat retrieval in EDTA at pH 8.0, but it does not establish whether retrieval is required across other preparations (caption: EDTA pH 8.0 retrieval). HPA has ICC-IF images in HaCaT and U2OS, but the supplied evidence does not establish whether frozen sections or IF are easier than paraffin IHC; bronchial mucus or inflammatory cells may complicate background assessment (HPA: ICC-IF image cell lines; standard IHC practice).
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, compartment patterns and controls before interpreting chromogenic signal.
What retrieval should I start with when STK3 staining is weak in paraffin sections?
Start with heat-mediated antigen retrieval in EDTA at pH 8.0 for paraffin sections (datasheet A02224-2). The selected image documents STK3 staining after that retrieval, followed by 10% goat serum block, 2 μg/mL primary antibody overnight at 4°C, and DAB detection (datasheet A02224-2). If staining remains weak, compare a second retrieval condition on adjacent sections while holding antibody concentration, detection and imaging constant (standard IHC practice). Include a retrieval-matched no-primary control and inspect tissue integrity, since stronger color alone cannot establish specific STK3 detection (standard IHC practice).
How should I troubleshoot variable STK3 staining between differently fixed specimens?
The selected tissue caption identifies a paraffin section but does not report its fixative, so target-specific fixation sensitivity is unknown (datasheet A02224-2). Record each specimen's fixative and fixation duration, then compare similarly processed sections using the documented EDTA pH 8.0 retrieval and 2 μg/mL primary concentration (datasheet A02224-2; standard IHC practice). Assess morphology and no-primary controls alongside signal, because fixation differences can alter antigen accessibility and background in IHC generally (standard IHC practice). Do not infer an STK3-specific fixation preference from its tissue staining pattern, topology or phosphorylation sites; test the actual specimens under matched conditions (standard IHC practice).
Should nuclear or filament-like STK3 staining be considered specific?
Assess cytoplasmic staining first: the tissue profile reports general cytoplasmic expression, while STK3 is also annotated in the nucleus and at centrosomes (HPA tissue IHC; UniProt Q13188 subcellular location). Nuclear signal can be biologically plausible because the caspase-cleaved form cycles between nucleus and cytoplasm, and Thr-117 phosphorylation inhibits nuclear translocation (UniProt Q13188 subcellular location). Intermediate-filament and rods-and-rings patterns are reported from cell imaging, but their appearance alone does not validate a chromogenic tissue pattern (HPA subcellular). Compare compartment scores with no-primary controls and morphology across matched sections before assigning a focal signal to STK3 (standard IHC practice).
Could epitope placement explain discordant STK3 staining across antibodies?
Map each antibody's disclosed immunogen or epitope against STK3 before comparing staining; the supplied caption does not specify the A02224-2 epitope (datasheet A02224-2). STK3 has 2 annotated isoforms, a kinase domain at residues 27–278, and a SARAH domain at 437–484 (UniProt Q13188 isoforms and domains). Processing yields annotated 1–322 and 323–491 chains, so antibodies against different regions could detect different processed forms (UniProt Q13188 processing; standard antibody interpretation). Because STK3 also carries phosphorylation sites, verify whether an antibody is modification-specific before treating staining differences as changes in total protein (UniProt Q13188 modified residues; standard IHC practice).
How should I adapt STK3 localisation checks to multiplex IF?
For a separate IF experiment, pair STK3 with a validated marker for the cell population being assessed, such as an adipocyte marker when examining adipocytes (HPA tissue IHC; standard IF practice). Assign the dimmer target to a bright, spectrally separated fluorophore and inspect unstained tissue for autofluorescence before choosing channels (standard IF practice). STK3 has no transmembrane segment and is annotated in cytoplasm and nucleus, so use a permeabilisation condition that permits access to intracellular epitopes while preserving the compartment being scored (UniProt Q13188 topology and subcellular location; standard IF practice). Include single-label and no-primary controls when judging overlap or punctate signal (standard IF practice).
What should I check when DAB staining appears diffuse or widespread?
Compare the primary-stained section with a no-primary section processed through the same secondary and DAB steps to locate detection-system background (standard IHC practice). The selected workflow uses a biotinylated goat anti-rabbit secondary and streptavidin-biotin complex, so assess background from that detection system separately from primary-antibody binding (datasheet A02224-2; standard IHC practice). Apply an appropriate endogenous peroxidase block and, where needed, a biotin-system control before interpreting brown signal (standard IHC practice). Broad cytoplasmic staining may still be plausible for STK3, so judge excess background against tissue morphology and matched controls rather than requiring a sharply restricted pattern (HPA tissue IHC; standard IHC practice).
How should I quantify STK3 staining across heterogeneous tissue sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, since STK3 is generally cytoplasmic in tissue IHC but also has annotated nuclear localisation (HPA tissue IHC; UniProt Q13188 subcellular location). For cell-level staining, report the percentage of positive cells and an intensity-weighted H-score using fixed thresholds across slides (standard IHC practice). If measuring positive-cell density, divide counts by analyzed viable tissue area in mm², and exclude folds, edges and necrotic regions consistently (standard IHC practice). Normalize comparisons to the same cell population, section-processing batch and detection settings, with control-section performance reported alongside the score (standard IHC practice).
How can I distinguish genuine STK3 signal from staining artefacts?
Look for reproducible cytoplasmic signal within intact cells and interpret nuclear staining cautiously in light of STK3's documented compartment cycling after caspase cleavage (HPA tissue IHC; UniProt Q13188 subcellular location). Compare cell identity with the scored population: HPA reports medium staining in adipocytes and several glandular or epithelial populations, but also low staining in listed squamous epithelial and hematopoietic populations (HPA tissue IHC). Flag color concentrated at section edges or necrotic areas, and check a no-primary section for endogenous enzyme or detection-system signal (standard IHC practice). Require consistent staining across comparable fields and controls before calling an unusual compartment or cell population positive (standard IHC practice).
Boster reagents
Best STK3 / Serine/threonine-protein kinase 3 IHC Antibodies
Anti-STK3 antibodies have IHC images from human paraffin sections of renal cancer tissue and kidney, plus an IF image from PC-3 cells (catalog image captions).
Real IHC data
Anti-STK3/MST-2 Antibody ®
Cat # A02224-2
Real IHC data
Anti-STK3/Mst2 Rabbit Monoclonal Antibody
Cat # M02224
A02224-2 has IHC data from human renal cancer paraffin sections and IF/ICC data from PC-3 cells (A02224-2 image captions). M02224 has IHC data from paraffin-embedded human kidney; its listed applications include IF/ICC, but no IF image is provided (M02224 catalog).
Which to pick: For tissue IHC, both SKUs have human paraffin-section images; A02224-2 provides a documented EDTA retrieval and 2 μg/ml staining example, while M02224 is a rabbit monoclonal with a kidney image (A02224-2 IHC caption; M02224 catalog and IHC caption). For IF/ICC, A02224-2 has a PC-3 image at 5 μg/ml; for mouse or rat samples, M02224 lists reactivity to those species, though its supplied IHC image is human (A02224-2 IF caption; M02224 catalog and IHC caption). Neither IHC caption reports the fixative (catalog IHC image captions).
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 fixation consistent across paraffin sections. (standard IHC practice; not target-specific); Caveat: Caspase cleavage can shift STK3 into nuclei (UniProt); Isoform / epitope: 2 isoforms; cleavage yields residues 1–322 and 323–491 , so check epitope placement (UniProt); Regulation: Caspase cleavage favors 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.