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JAK2 / Tyrosine-protein kinase JAK2 · IHC design guide
Design Immunohistochemistry for JAK2
Plan JAK2 paraffin IHC around cytoplasmic staining, with strong signal in lung macrophages and gallbladder glandular cells (HPA tissue IHC). This guide covers the catalog antibody’s 1:50 dilution, consistent fixation, chromogenic detection, and scoring by cell type (datasheet: M00027; standard IHC practice).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
JAK2 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 staining in several tissues (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic staining, strongest in subsets of immune cells (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet M00027) | |
| Positive control | Gallbladder+4 more · see all | |
| Negative control | Caudate+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent across sections (standard IHC practice; not target-specific) | |
| Caveat | Immune-cell-rich regions may dominate the signal (HPA tissue IHC) | |
| Regulation | Expression regulation not specified (UniProt) | |
| Isoform / epitope | 0 annotated isoforms; no processing or membrane span (UniProt) |
Section 1
Recommended JAK2 IHC & IF Protocols
Compare the catalog antibody’s IHC-P protocol (datasheet M00027) with four published JAK2 IHC workflows; one measures phospho-JAK2 (PMC4084704).
| Sample | Paraffin-embedded human prostate cancer tissue; fixative not specified (datasheet M00027) |
| Fixation | Image fixative and duration unreported (datasheet M00027); 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 M00027); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet M00027) |
| Primary antibody | Rabbit monoclonal (clone COG-10) anti-JAK2, 1:50 (datasheet M00027) |
| Primary incubation | Overnight at 4 °C (datasheet M00027) |
| Detection | HRP-conjugated secondary, DAB chromogen (datasheet M00027) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | JAK2-positive staining in glandular cells of gallbladder (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in several tissues, most abundant in subsets of immune cells. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA retrieval at pH 8.0 (datasheet M00027); the bone marrow protocol also uses microwave EDTA at pH 8.0 (PMC7433821).
Section 2
What Is the Expected JAK2 Staining Pattern?
In paraffin-section IHC, expect predominantly cytoplasmic JAK2 staining, especially in subsets of immune cells (HPA: tissue IHC). Lung macrophages and lymph-node and tonsil non-germinal center cells show high staining (HPA: High). JAK2 has no transmembrane segment, although it participates in receptor signaling and is also annotated in the nucleus (UniProt O60674: topology, function, subcellular location). HPA rates the tissue IHC pattern Approved, with medium consistency against RNA data (HPA: reliability).
| Cytoplasmic staining is strong in lung macrophages or non-germinal center cells of lymph node or tonsil. | This matches the reported high-staining cell populations and provides a practical positive-pattern check; it does not imply that every cell in the section should stain (HPA: tissue IHC). |
| An IHC section shows predominantly nuclear staining with little cytoplasmic staining in expected positive cells. | Recheck specificity and detection before calling it the expected tissue pattern: HPA describes tissue IHC as cytoplasmic, while UniProt also lists nuclear localization (HPA: tissue IHC; UniProt O60674: subcellular location). |
| Strong staining appears mainly in cell types listed as not detected, such as skeletal-muscle myocytes. | Treat this as a specificity or detection concern, not proof that JAK2 is absent there: HPA reports a staining observation, and its tissue IHC reliability has medium RNA consistency (HPA: Not detected in myocytes; reliability). |
| Brown deposit spreads across tissue and obscures cell boundaries. | The pattern cannot be scored confidently by cell type or compartment. Check a no-primary control for detection background, then compare staining with the cytoplasmic, cell-restricted HPA pattern (general IHC practice; HPA: tissue IHC). |
| No stain is visible in an included lung macrophage-rich area. | A technical failure is possible because HPA reports high staining in lung macrophages. First establish that the expected cells are present and the detection run worked; an individual negative section is not decisive (HPA: High in lung macrophages; general IHC practice). |
💡Expected JAK2 appearanceCall the result positive when staining is predominantly cytoplasmic in the expected cells, with high signal in lung macrophages or lymphoid non-germinal center cells; widespread deposit or dominant nuclear-only IHC staining warrants review (HPA: tissue IHC; HPA: High in named cells).
| Tissue and cell selection | JAK2 is broadly expressed, but IHC intensity varies by cell population; use the named high-staining cells as pattern references and interpret listed not-detected cells at the cell level (UniProt O60674: tissue specificity; HPA: tissue IHC). |
| Membrane-associated signal | JAK2 lacks a transmembrane segment and acts with receptors through their intracellular signaling machinery. A crisp membrane-only IHC pattern should therefore be checked against the reported cytoplasmic tissue pattern (UniProt O60674: topology, function; HPA: tissue IHC). |
| Antibody and epitope scope | HPA lists two IHC antibodies as Approved, but the supplied record gives no epitope or phospho-specificity for the antibody used here. JAK2 has annotated phosphorylated residues, so staining alone cannot establish activation state (HPA: antibody validation; UniProt O60674: modified residues). |
| IF/ICC Q&A: why might nuclear staining appear? | HPA reports supported nucleoplasmic localization in ICC-IF, whereas its tissue IHC profile is cytoplasmic. Interpret each assay against its own evidence; nuclear IF signal does not make nuclear-only chromogenic IHC an established tissue pattern (HPA: ICC-IF subcellular; tissue IHC). |
| Situation | Likely cause | Next action |
|---|---|---|
| Expected high-staining cells are present, but IHC signal is absent. | The run may lack sensitivity or have a detection problem; the supplied sources give no JAK2-specific retrieval condition (HPA: High in lung macrophages and lymphoid non-germinal center cells). | Check the run controls and detection steps; optimize retrieval and antibody concentration within the validated IHC workflow, then reassess the expected cells (general IHC practice). |
| Staining is widespread and poorly confined to cells. | Detection background may obscure the reported cytoplasmic pattern; HPA tissue observations alone cannot identify the chemical source of that background (HPA: tissue IHC). | Review the no-primary control, blocking and washes, and compare the signal with the expected cellular distribution (general IHC practice; HPA: tissue IHC). |
| Macrophage-rich areas show granular brown signal even without primary antibody. | Endogenous detection activity is a possible source of chromogenic background in such areas (general IHC practice). | Use the no-primary control to distinguish detection activity from antibody-dependent stain; review the detection-system blocking step before scoring macrophages (general IHC practice). |
| Nuclear staining dominates a paraffin section. | This differs from HPA's cytoplasmic tissue IHC profile, although nuclear JAK2 is annotated and nucleoplasmic ICC-IF is supported (HPA: tissue IHC, ICC-IF; UniProt O60674: subcellular location). | Compare cytoplasmic signal in expected positive cells and review controls before accepting a nuclear IHC interpretation; keep the IF observation assay-specific (HPA: tissue IHC, ICC-IF; general IHC practice). |
| Cells listed as not detected stain strongly. | Cross-reactivity or detection background is possible; a single discordant section cannot resolve specificity (HPA: Not detected in listed cell types; reliability). | Check the no-primary control and repeat the comparison alongside an HPA high-staining cell population using the same IHC conditions (HPA: tissue IHC; general IHC practice). |
| Staining intensity varies among cells within one tissue. | This can reflect the cell-specific distribution: HPA reports high, medium, low and not-detected staining across named populations (HPA: tissue IHC). | Score the identified cell type and its cytoplasmic pattern separately from nearby cells; avoid treating whole-section intensity as a single JAK2 result (HPA: tissue IHC; general IHC practice). |
Experiment evidence
IHC & IF images showing JAK2 staining
M00027 · IHCIHC analysis of JAK2 using anti-JAK2 antibody (M00027). JAK2 was detected in a paraffin-embedded section of human prostate cancer tissue. He
M00027 · IFIF analysis of JAK2 using anti-JAK2 antibody (M00027) and anti-Beta Tubulin antibody (M01857-3). JAK2 was detected in immunocytochemical sec
M00027-3 · IHCHuman lung carcinoma was stained with anti-JAK2 rabbit antibody
M00027-3 · IHCMouse lung was stained with anti-JAK2 rabbit antibody
MP00027 · IHCImmunohistochemical analysis of paraffin-embedded human cervix cancer, using Phospho-JAK2 (Y1007 + Y1008) Antibody.
MP00027 · IFImmunofluorescent analysis of Jurkat cells treated with Pervanadate, using Phospho-JAK2 (Y1007 + Y1008) Antibody
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000096968-JAK2/tissue/cerebral+cortex#imid_3351516
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000096968-JAK2/tissue/colon#imid_3351458
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000096968-JAK2/tissue/liver#imid_3351451
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000096968-JAK2/tissue/kidney#imid_3351460
HPA · ICC-IF HeLaHuman Protein Atlas immunofluorescence, HeLa, antibody HPA043870; green = JAK2, blue = nucleus, red = microtubules.
HPA · ICC-IF HeLaHuman Protein Atlas immunofluorescence, HeLa, antibody HPA043870; green = JAK2, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for JAK2 IHC & IF
🧪Run gallbladder first and expect staining in its glandular cells (HPA: High in gallbladder glandular cells). Use cervix glandular cells as a negative comparator (HPA: Not detected in cervix glandular cells); on the gallbladder slide, treat any unstained adjacent cells as background references, without assuming they are JAK2-negative (HPA: gallbladder glandular-cell result only).
Positive control tissue: Gallbladder (Glandular cells, HPA High)
Negative control tissue: Caudate (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show JAK2 in HeLa, Rh30, U2OS, hTERT-RPE1 (serum starved), KOLF2.1J, with annotated localisation: Nucleoplasm (supported) (HPA subcellular).
Technical controls: Include a no-primary (secondary-only) control, a rabbit IgG isotype control matched to the primary’s clonality when known, and identically processed JAK2-knockout material if available (caption: rabbit anti-JAK2; standard IHC practice). For gallbladder HRP/DAB staining, quench endogenous peroxidase and check for bile pigment that could resemble DAB; block endogenous biotin if using avidin–biotin detection (caption: HRP/DAB; standard IHC practice).
⚠️Feasibility: A target-specific fixation window and fixation effect are unreported, and the selected M00027 paraffin-section caption does not state the fixative (caption: fixative unreported). The caption uses EDTA retrieval at pH 8.0, but does not establish that retrieval is required or whether frozen sections or IF are easier (caption: EDTA retrieval; no matched comparison supplied). In gallbladder, assess bile pigment against the no-primary control when scoring brown DAB signal (standard IHC practice).
HPA tissue IHC evidence for JAK2
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 |
|---|---|---|---|---|
| Gallbladder | Glandular cells | High | Protein (IHC) | HPA → |
| Lung | Macrophages | High | Protein (IHC) | HPA → |
| Lymph node | Non-germinal center cells | High | Protein (IHC) | HPA → |
| Tonsil | Non-germinal center cells | High | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | Medium | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Caudate | Glial cells | Not detected | Protein (IHC) | HPA → |
| Cervix | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Esophagus | Squamous epithelial cells | Not detected | Protein (IHC) | HPA → |
| Prostate | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Skeletal muscle | Myocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced JAK2 IHC Tips
Troubleshoot JAK2 staining in paraffin sections by checking retrieval, compartment, cell type and detection controls against the available tissue evidence.
What retrieval should I start with for weak JAK2 staining in paraffin sections?
Start with heat-mediated retrieval in EDTA at pH 8.0 for paraffin sections (datasheet M00027). The documented prostate cancer section used that retrieval before 10% goat-serum blocking and an overnight 1:50 primary incubation at 4°C (datasheet M00027). If staining is weak, compare a longer or shorter heating period on serial sections while holding antibody concentration and detection constant (standard IHC practice). Evaluate cytoplasmic signal in relevant cells alongside tissue morphology; the tissue survey reports cytoplasmic JAK2, especially in subsets of immune cells (HPA tissue IHC). Record retrieval temperature, heating time and cooling time so a stronger signal can be distinguished from damaged morphology (standard IHC practice).
How should I troubleshoot possible fixation effects on JAK2 IHC?
JAK2-specific sensitivity to fixation is unknown from the supplied evidence; the catalog image identifies a paraffin section but does not state its fixative (datasheet M00027). Record the fixative, fixation duration and processing history for each specimen before comparing staining across cases (standard IHC practice). On matched serial sections, keep retrieval at EDTA pH 8.0 and the detection workflow constant while assessing whether staining tracks with tissue preservation (datasheet M00027; standard IHC practice). Compare suspect areas with well-preserved areas and a concurrently processed positive control, watching for uneven staining at section edges (standard IHC practice). Do not infer a JAK2-specific fixation effect from its localisation or phosphorylation sites (UniProt O60674).
Which compartments should show JAK2, and how should unexpected nuclear staining be assessed?
In tissue IHC, cytoplasmic staining is the main reported pattern, with abundant signal in subsets of immune cells (HPA tissue IHC). Nuclear staining is biologically plausible because JAK2 is annotated in the nucleus and can phosphorylate histone H3, while ICC/IF places its main signal in the nucleoplasm (UniProt O60674; HPA subcellular). JAK2 has no transmembrane segment, so a crisp membrane-only pattern needs independent support even though receptor association is part of its biology (UniProt O60674). Score cytoplasmic and nuclear compartments separately, checking morphology and a no-primary control before accepting an unusual pattern (standard IHC practice). Treat plasma-membrane ICC/IF localisation cautiously because its HPA designation is uncertain (HPA subcellular).
How can epitope placement affect interpretation of JAK2 staining?
The supplied record lists 0 isoforms and one 1–1132 chain, but it does not give the catalog antibody’s epitope (UniProt O60674; datasheet M00027). JAK2 contains a FERM region at residues 37–380 and kinase regions at 545–809 and 849–1124, so epitope location would matter when interpreting altered protein forms (UniProt O60674). Its annotated phosphorylation sites include tyrosines 119, 372 and 373, but those annotations do not establish phospho-specific recognition by this antibody (UniProt O60674). Check the antibody’s epitope documentation before attributing compartment differences to a particular domain or modification (standard IHC practice). If specificity remains uncertain, compare staining with an independently validated JAK2 reagent on matched sections (standard IHC practice).
How can IF help assess an ambiguous JAK2 IHC pattern?
Use IF as a complementary localisation check rather than a substitute for the documented paraffin-section chromogenic workflow (datasheet M00027; standard IHC/IF practice). Multiplex JAK2 with a validated marker for the expected cell population; lung macrophages show high JAK2 tissue staining and provide a cell-type reference (HPA tissue IHC). Choose fluorophores whose emission is well separated from the specimen’s autofluorescence, and inspect unstained tissue in each channel (standard IF practice). For intracellular epitopes, optimise permeabilisation to reach cytoplasm or nucleoplasm; JAK2 lacks a transmembrane segment, and its ICC/IF main location is nucleoplasmic (UniProt O60674; HPA subcellular). Assess channel bleed-through and secondary-only controls before comparing IF with IHC (standard IF practice).
What should I change when JAK2 DAB staining is diffuse or widespread?
First compare a no-primary section with the test section to identify signal arising from detection reagents or endogenous peroxidase (standard IHC practice). The documented workflow used 10% goat-serum blocking, a 1:50 primary incubation overnight at 4°C, an HRP secondary and DAB (datasheet M00027). If background persists, optimise the general peroxidase block, shorten DAB development and titrate the primary on serial sections while preserving a positive control (standard IHC practice). Distinguish widespread low-level colour from cell-specific cytoplasmic staining, particularly in immune-cell subsets where JAK2 staining is abundant (HPA tissue IHC). Document each adjustment independently so loss of background can be weighed against loss of expected signal (standard IHC practice).
How should I quantify JAK2 across sections with different cell composition? ⚠ ANSWER MARKED FOR VERIFICATION
Define the compartment and cell population before scoring; the tissue survey reports cytoplasmic expression, with high staining in lung macrophages and several other specified cell populations (HPA tissue IHC). For a defined cell population, report an H-score or the percentage of positive cells using one staining threshold across the batch (standard IHC practice). If the question concerns infiltrating cells, report positive-cell density per mm² of viable tissue and normalise it to the density of that cell population (standard IHC practice). Score nuclear and cytoplasmic signal separately when both occur, because nuclear localisation is also supported by independent subcellular evidence (UniProt O60674; HPA subcellular). Exclude folds, edges and necrosis using prespecified rules (standard IHC practice).
How do I distinguish convincing JAK2 positivity from staining artefacts?
Look for reproducible cellular staining in morphologically intact regions, with cytoplasmic signal fitting the tissue IHC profile and the sampled cell type (HPA tissue IHC; standard IHC practice). Nuclear signal can be plausible, but an isolated compartment pattern should be checked against controls because JAK2’s nuclear annotation and nucleoplasmic ICC/IF localisation do not validate every tissue-IHC nucleus (UniProt O60674; HPA subcellular). Compare suspicious areas with adjacent viable tissue and a no-primary section; edge enhancement, necrotic deposits and endogenous peroxidase can mimic DAB positivity (standard IHC practice). Consider the precise cell population: normal prostate glandular cells were reported as not detected, whereas the catalog image shows staining in prostate cancer tissue (HPA tissue IHC; datasheet M00027). Resolve discordant patterns with repeat sections and an independent antibody if available (standard IHC practice).
Boster reagents
Best JAK2 / Tyrosine-protein kinase JAK2 IHC Antibodies
The catalog includes JAK2 antibodies with IHC images from human, mouse, and rat tissues (catalog IHC captions) and IF images from human cells (catalog IF captions).
Real IHC data
Anti-JAK2 Monoclonal Antibody
Cat # M00027
Real IHC data
Anti-JAK2 Rabbit Monoclonal Antibody
Cat # M00027-3
Real IHC data
Anti-Phospho-JAK2 (Y1007 + Y1008) Monoclonal Antibody
Cat # MP00027
M00027 has IHC data from a paraffin-embedded human prostate cancer section and IF data from A549 cells (M00027 image captions). M00027-3 has IHC images from human lung carcinoma, mouse lung, and rat lung; MP00027 has phospho-JAK2 IHC data from paraffin-embedded human cervix cancer and IF data from treated Jurkat cells (respective image captions).
Which to pick: For tissue IHC, choose M00027 when its documented paraffin-section procedure is useful: EDTA retrieval at pH 8.0 and a 1:50 primary dilution (M00027 IHC caption); the fixative is unreported (M00027 IHC caption). For IF/ICC, M00027 has an A549 IF example and lists both IF and ICC, while MP00027 targets phospho-JAK2 Y1007/Y1008 and has a Jurkat IF example (catalog applications and IF captions). For cross-species tissue IHC, M00027-3 has human, mouse, and rat lung images, although their captions do not specify tissue processing or fixative (M00027-3 IHC captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry O60674 (JAK2_HUMAN, Tyrosine-protein kinase JAK2).
- Human Protein Atlas. JAK2 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. JAK2 subcellular location (ICC-IF): Mainly localized to the nucleoplasm. In addition localized to the plasma membrane and focal adhesion sites..
- Human Protein Atlas. JAK2 antibody validation summary (3 antibodies).
- The impact of bone marrow fibrosis and JAK2 expression on clinical outcomes in patients with newly diagnosed multiple myeloma treated with immunomodulatory agents and/or proteasome inhibitors. Cancer medicine 2020 — PMC7433821.
- Selective JAK2 inhibition specifically decreases Hodgkin lymphoma and mediastinal large B-cell lymphoma growth in vitro and in vivo. Clinical cancer research : an official journal of the American Association for Cancer Research 2014 — PMC4084704.
- Proinflammatory State in the Odontogenesis of Fetuses Exposed to Different Types of Fatty Acids during Pregnancy. Medical principles and practice : international journal of the Kuwait University, Health Science Centre 2022 — PMC9841762.
- The impact of tocilizumab treatment on the severity of inflammation and survival rates in sepsis is significantly influence by the timing of administration. Inflammopharmacology 2025 — PMC11914034.
- PubMed PMID:9618263 — UniProt-cited evidence.
- PubMed PMID:9446644 — UniProt-cited evidence.
- PubMed PMID:9326218 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for JAK2 (Tyrosine-protein kinase JAK2). Expected localisation Cytoplasmic staining in several tissues (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet M00027). Validated IHC antibody: M00027 at 1:50 (the product's own IHC validation image is shown). Controls (HPA): positive Gallbladder (Glandular cells); negative Caudate. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human prostate cancer tissue; fixative not specified (datasheet M00027); 2. Fixation: Image fixative and duration unreported (datasheet M00027); 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 M00027); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet M00027); 8. Primary antibody: Rabbit monoclonal (clone COG-10) anti-JAK2, 1:50 (datasheet M00027); 9. Primary incubation: Overnight at 4 °C (datasheet M00027). Watch for: Fixation: Keep fixation consistent across sections (standard IHC practice; not target-specific); Caveat: Immune-cell-rich regions may dominate the signal (HPA tissue IHC); Isoform / epitope: 0 annotated isoforms; no processing or membrane span (UniProt); Regulation: Expression regulation not specified (UniProt). Evidence: UniProt O60674, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the nucleoplasm. In addition localized to the…, Datasheet M00027, PMC7433821, PMC4084704. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/JAK2/immunohistochemistry-ihc-guide.