Phospho-JAK2 (Y1007 + Y1008) Recombinant Monoclonal Antibody
Description
Mechanism of Action and Significance
Phosphorylation at Y1007 and Y1008 in JAK2’s activation loop is essential for its kinase activity. This modification enables downstream signaling via the JAK/STAT pathway, regulating processes like hematopoiesis and immune responses .
Key Research Findings
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Regulation of JAK2 Activity:
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Phosphorylation at Y972 (a regulatory site) is required for Y1007/Y1008 phosphorylation. Mutating Y972 to phenylalanine reduces total JAK2 tyrosine phosphorylation and Y1007/Y1008-specific phosphorylation .
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CHZ868 (a type II JAK2 inhibitor) inhibits Y1007/Y1008 phosphorylation, reversing persistent activation caused by type I inhibitors like ruxolitinib .
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Cross-Reactivity and Specificity
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TYK2 and JAK1: The antibody may cross-react with phosphorylated TYK2 or JAK1 due to sequence homology in activation loops .
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Validation: Specificity is confirmed via Western blot and immunoprecipitation assays, often using phospho-peptide competitors or JAK2-depleted cell lines .
Applications in Research and Diagnostics
The antibody is widely used to study JAK2 signaling in diseases like myeloproliferative neoplasms (MPNs) and leukemia.
Experimental Uses
Case Study: In Jurkat T-cells, treatment with pervanadate induces robust Y1007/Y1008 phosphorylation, detectable via WB and flow cytometry .
Clinical and Therapeutic Relevance
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Disease Biomarker: Phospho-JAK2 (Y1007/Y1008) levels correlate with disease progression in MPNs and JAK2 V617F-driven leukemias .
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Drug Development: The antibody is used to assess JAK inhibitor efficacy, such as CHZ868’s ability to suppress Y1007/Y1008 phosphorylation in MPLW515L mutant cells .
Western Blot Protocol
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Lysis: Use RIPA buffer with phosphatase inhibitors (e.g., Na3VO4).
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Blocking: 5% BSA/TBST for 1 hour at RT.
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Primary Antibody: Incubate overnight at 4°C (1:500–1:5000 dilution) .
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Detection: HRP-conjugated secondary antibody (1:20,000) + ECL substrate .
Flow Cytometry Protocol
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Fixation: 4% PFA for 10 minutes.
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Permeabilization: 90% methanol for 30 minutes.
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Staining: Incubate with antibody (1:20–1:50) for 1 hour at RT .
Limitations and Considerations
Product Specs
This rabbit IgG recombinant monoclonal antibody specifically recognizes human JAK2 phosphorylated at tyrosine residues 1007 and 1008. The antibody's DNA sequence was cloned into a plasmid vector, transfected into a suitable cell line for expression, and purified using affinity chromatography. This recombinant phospho-JAK2 (Y1007 + Y1008) antibody exhibits reactivity with human samples and has been validated for use in ELISA, Western blotting (WB), immunohistochemistry (IHC), and immunoprecipitation (IP).
JAK2 is a non-receptor protein tyrosine kinase belonging to the Janus kinase (JAK) family, which also includes JAK1, JAK3, and TYK2. JAKs are cytoplasmic signaling molecules associated with cytokine receptors. Cytokine binding triggers trans-phosphorylation, leading to receptor and downstream STAT protein phosphorylation. Upon cytokine stimulation, JAK2 undergoes phosphorylation at multiple sites, including Tyr1007 and Tyr1008.
Target Background
JAK2, a non-receptor tyrosine kinase, plays a critical role in diverse cellular processes, including growth, development, differentiation, and histone modification. It is a pivotal signaling component in both innate and adaptive immunity. In the cytoplasm, JAK2 associates with type I receptors (e.g., growth hormone receptor (GHR), prolactin receptor (PRLR), leptin receptor (LEPR), erythropoietin receptor (EPOR), thrombopoietin receptor (THPO)) and type II receptors (e.g., IFN-α, IFN-β, IFN-γ, and various interleukin receptors). Ligand binding to these receptors leads to JAK2 autophosphorylation and subsequent receptor phosphorylation, creating docking sites for STAT proteins. JAK2 then phosphorylates the recruited STAT proteins, which dimerize and translocate to the nucleus to regulate gene transcription. For instance, in erythropoiesis, erythropoietin (EPO) stimulation causes JAK2 autophosphorylation and association with the EPOR, resulting in EPOR phosphorylation and subsequent STAT5 (STAT5A or STAT5B) recruitment, phosphorylation, activation, and nuclear translocation to modulate gene expression essential for erythropoiesis. JAK2 is also involved in a signaling cascade activated by increased cellular retinol, leading to STAT5 activation. Additionally, JAK2 mediates angiotensin-II-induced ARHGEF1 phosphorylation, regulates the cell cycle by phosphorylating CDKN1B, and cooperates with TEC through reciprocal phosphorylation to modulate cytokine-driven FOS transcription. Within the nucleus, JAK2 phosphorylates histone H3 at tyrosine 41 (H3Y41ph), a modification implicated in the exclusion of CBX5 (HP1α) from chromatin.
- Clonal analysis reveals that the predominant JAK2 V617F-positive clone in polycythemia vera often harbors an EGFR C329R substitution, suggesting a contribution to clonal expansion. PMID: 28550306
- Patients with CALR mutations exhibit significantly higher PDGF-BB and lower SDF-1α concentrations compared to those with JAK2V617F mutations. Elevated PDGF-BB and reduced SDF-1α levels in CALR(+) essential thrombocythemia (ET) may indicate a role for these chemokines in platelet calcium metabolism dysregulation. PMID: 29390868
- Crystal structures of human JAK2 FERM and SH2 domains bound to LEPR and EPOR reveal a novel JAK2 dimeric conformation. PMID: 30044226
- Investigation into the pathogenesis of the JAK2 F556V mutation in myeloproliferative neoplasms (MPNs). PMID: 29842959
- miR-204 attenuates angiogenesis in lung adenocarcinoma via the JAK2-STAT3 pathway. PMID: 29281186
- FEZF1-AS1 acts as an oncogenic long non-coding RNA (lncRNA) in hepatocellular carcinoma by promoting epithelial-mesenchymal transition (EMT) through JAK2/STAT3 signaling. PMID: 29957463
- Case reports and review of JAK2 mutation-associated cerebral arterial infarction and cerebral and systemic venous thromboembolism. PMID: 30056970
- HSP27 interacts with JAK2-STAT5 and is a potential therapeutic target in myelofibrosis. PMID: 29650953
- The JAK2V617F mutation may increase the risk of thrombosis in chronic myeloproliferative neoplasms. PMID: 30004057
- Progression to polycythemia vera from familial thrombocytosis with a germline JAK2 R867Q mutation. PMID: 29368262
- JAK2 and STAT3 activation in idiopathic pulmonary fibrosis. PMID: 29409529
- The prevalence of CALR mutations in JAK2V617F-negative essential thrombocythemia is approximately 35.7%. High-resolution melting (HRM) analysis is an effective method for detecting CALR mutations. PMID: 29521158
- Genomic characterization of myeloproliferative neoplasms identified distinct genetic subgroups and revealed that mutations in JAK2, CALR, or MPL are the sole abnormality in 45% of patients. PMID: 30304655
- Inhibition of P16 reduces breast cancer cell growth and metastasis by inhibiting IL-6/JAK2/STAT3 signaling. PMID: 29388151
- MPL-mutated and CALR-mutated essential thrombocythemia share clinical and histological characteristics, with both showing higher platelet counts and marked megakaryocytic proliferation compared to JAK2V617F-mutated ET. PMID: 29934356
- Insights into the mechanism by which the JAK2 V625F mutation causes myeloproliferative neoplasms, informing the development of JAK2 mutation-specific inhibitors. PMID: 29782975
- Concomitant presence of JAK2V617F mutation and BCR-ABL translocation in two patients: a novel entity or a variant of myeloproliferative neoplasms. PMID: 29845291
- The JAK2 V617F mutation and thrombocytopenia. PMID: 27614229
- PBX1 plays an oncogenic role in clear cell renal carcinoma via the JAK2/STAT3 pathway. PMID: 29678569
- JAK2V617F leads to abnormal expression of membrane proteins in polycythemia vera red blood cells, including CALR overexpression and CANX persistence. PMID: 28385780
- Mutations in JAK2, MPL, or CALR are found in 94.9% of polycythemia vera, 85.5% of essential thrombocythemia, and 85.2% of primary myelofibrosis cases. PMID: 28990497
- Tyrphostin B42 induces apoptosis in pancreatic cancer cells by regulating mitochondrial genes and antagonizes trichostatin A resistance by inhibiting IL6/JAK2/STAT3 signaling. PMID: 29393444
- miR-375 inhibits fetal airway smooth muscle cell proliferation and migration by targeting JAK2/STAT3 signaling. PMID: 29245068
- Heparin-induced thrombocytopenia (HIT) is more frequent during heparin treatment in patients with ET carrying the V617F mutation compared to those without mutations. PMID: 29022213
- ALK4 overexpression suppresses glioma cell proliferation, migration, and invasion through inactivation of the JAK/STAT3 signaling pathway. PMID: 29278854
- A subset of non-small-cell lung cancer patients exhibit JAK2 amplifications resulting in high PD-L1 expression. PMID: 28795418
- High JAK2 expression is associated with hepatocellular carcinoma. PMID: 28677802
- JAK2 haplotype 46/1 and JAK2 V617F allele burden in MPNs. PMID: 29134760
- Low JAK2 expression is associated with gastric cancer. PMID: 28656307
- Tyrosine 78 of Atoh1 is phosphorylated by a Jak2-mediated pathway in tumor-initiating cells and human Sonic Hedgehog-type medulloblastoma. PMID: 29168692
- The activating JAK2 V617F mutation does not play a decisive role in the pathogenesis of progressive chronic kidney disease (CKD). PMID: 27889755
- B7-H3 affects ovarian cancer progression through the Jak2/Stat3 pathway, suggesting its potential as a prognostic marker. PMID: 28765941
- In myelofibrosis patients undergoing allogeneic stem cell transplantation, molecular clearance on day 100 was higher in CALR-mutated patients (92%) compared to MPL- (75%) and JAKV617F-mutated patients (67%). PMID: 28714945
- JAK2 mutational subtypes correlate with different clinical features in Japanese patients with myeloproliferative neoplasms. PMID: 29464483
- Identification of activating somatic mutations in JAK2 and germline mutations in JAK3 with clinical implications. PMID: 29082853
- Screening for the JAK2 V617F mutation in cerebral venous thrombosis patients is useful due to its relatively high prevalence and risk of thrombosis recurrence. PMID: 28609766
- Ascochlorin significantly decreased JAK2/STAT3 phosphorylation, cancer cell migration, and nuclear STAT3 translocation. PMID: 28569433
- TLR7, TLR9, and JAK2 genes are potential biomarkers for systemic sclerosis. PMID: 29147913
- The JAK2V617F mutation is detectable in patients with stroke. PMID: 28625126
- Curcumin attenuated neuropathic pain and down-regulated spinal mature IL-1β production by inhibiting NALP1 inflammasome aggregation and JAK2-STAT3 cascade activation in astrocytes. PMID: 27381056
- High levels of phosphorylated JAK2 and STAT3 are associated with systemic lupus erythematosus. PMID: 28177455
- Nrf2 activation induces a lipocyte phenotype in hepatic stellate cells by enhancing SOCS3-dependent feedback inhibition on the JAK2/STAT3 cascade. PMID: 28601022
- Bladder cancer cells may inhibit dendritic cell maturation and function through the Jak2/STAT3 pathway, with potential differences in mechanisms for adriamycin-resistant bladder cancer cells. PMID: 27556503
- Increased activated B cells are universally present in JAK2-mutated, CALR-mutated, and triple-negative ET patients compared to healthy adults. PMID: 28415571
- Younger age, platelet count, hemoglobin level, and JAK2 V617F mutation independently predicted acquired von Willebrand syndrome (AVWS) development in essential thrombocythemia patients; platelet count was the main predictor in polycythemia vera patients. In ET patients, JAK2 V617F was a major driver of AVWS. PMID: 27919526
- CXCR4 induced VEGF production and JAK2/STAT3 activation, enhancing STAT3 binding to the VEGF promoter in gastric cancer cells. PMID: 28544312
- Proteome alterations in MPN granulocytes depend on patient phenotype and genotype, highlighting new oncogenic mechanisms associated with JAK2 mutations and calreticulin overexpression. PMID: 28314843
- JAK2 mutation is associated with essential thrombocythemia. PMID: 28205126
- In JAK2(V617F)-positive disease, a higher JAK2(V617F) burden (>50%) and histological classification are independent prognostic risk factors for disease progression. PMID: 28509339
- Silibinin inhibits the Jak2/STAT3/MMP2 signaling pathway and inhibits the proliferation, migration, and invasion of triple-negative breast cancer cells. PMID: 28440514
Q&A
Basic Research Questions
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What is the specificity of Phospho-JAK2 (Y1007 + Y1008) Monoclonal Antibody?
The Phospho-JAK2 (Y1007 + Y1008) Monoclonal Antibody specifically recognizes JAK2 that is phosphorylated at tyrosine residues 1007 and 1008 in the activation loop. The antibody is designed to detect activated forms of JAK2, including JAK2V617F mutant, with high specificity. It demonstrates minimal response to wild-type non-phosphorylated JAK2 and does not cross-react with other tyrosine kinases such as epidermal growth factor receptor or insulin receptor . The antibody's specificity makes it particularly valuable for distinguishing between inactive and active JAK2 in experimental settings.
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Which species does the Phospho-JAK2 (Y1007 + Y1008) Monoclonal Antibody react with?
According to validation data, the antibody has confirmed reactivity with human, mouse, and rat samples . While not explicitly tested for all species, researchers have inquired about cross-reactivity with pig tissues, which may be possible due to the conservation of JAK2 phosphorylation sites across mammals . When planning experiments with non-validated species, researchers should perform preliminary validation studies or utilize the supplier's innovator award programs that support such validation efforts.
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What applications is the Phospho-JAK2 (Y1007 + Y1008) Monoclonal Antibody validated for?
The antibody has been validated for multiple applications including Western Blotting (WB), Immunohistochemistry (IHC), Immunocytochemistry (ICC), Immunofluorescence (IF), Immunoprecipitation (IP), and Flow Cytometry . It has been successfully used for detection of phosphorylated JAK2 in various sample types including cell lines, primary tissues, and mononuclear cells from patient samples. The antibody has demonstrated particular efficacy in detecting JAK2V617F activity in polycythemia vera blood samples and is suitable for both qualitative and quantitative assays of JAK2 activity .
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What expression patterns of phosphorylated JAK2 have been documented in different tissues?
According to research data, phosphorylated JAK2 has been detected in several tissue types using the Phospho-JAK2 (Y1007 + Y1008) Monoclonal Antibody. Expression has been confirmed in erythroleukemia cells, where staining is observed in the endomembrane system . JAK2 expression has also been documented in fetal brain tissue, making these samples appropriate for research applications using this antibody . Additional tissues with JAK2 expression include popliteal artery and various hematopoietic cell lineages. When studying novel tissue types, researchers should include appropriate positive controls based on these known expression patterns.
