Replaces Prod. #: BML-UW8995
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Efficient tool for the selective isolation of ubiquitinylated proteins
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High binding affinity matrix, able to bind both monoubiquitinated and polyubiquitinated proteins
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Highly compatible with a wide range of lysate buffer, different sample types (cells, tissues) and a broad range of species
For isolation and enrichment of ubiquitinylated proteins.The UBI-QAPTURE-Q® kit is an efficient tool for the selective isolation of ubiquitinylated proteins. The Kit facilitates the isolation of both mono- and poly-ubiquitinylated proteins (independent of lysine residue chain linkage) from cell extracts, tissue lysates and in vitro assay solutions through use of a high-binding affinity matrix. Captured proteins can be analyzed by Western blotting using the highly sensitive ubiquitin-conjugate specific antibody provided, using antibodies to specific proteins of interest, or eluted from the matrix for subsequent biochemical characterization. The UBI-QAPTURE-Q® matrix supplied with the kit has superior binding characteristics compared to other commercially available matrices and is compatible with a wide range of lysate buffers and cell/tissue samples from a variety of species.
Western Blot analysis of ubiquitin enrichment of lysate derived ubiquitinylated proteins. Ubiquitin enrichment experiment set-up and run as described in UBIQAPTURE-Q® kit (Prod. No. BML-UW8995A). Ubiquitin-protein conjugates present in starting material, unbound fraction and elution fraction were detected by western blotting using the provided ubiquitin-conjugate specific HRP-linked antibody (Prod. No. ENZ-ABS840HRP) at a dilution of 1:1000. Key: SM = Starting Material, UF = Unbound Fraction, WA = Wash Fraction, and EL = Elution Fraction.
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Product Details
| Application Notes: | Uses:
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Isolation and detection of the full range of ubiquitinylated protein conjugates (mono-/multi-/poly-ubiquitin modified, lysine linkage independent) from specific cell/tissue lysates of interest, especially low abundance (regulatory) proteins involved in the ubiquitin-proteasome pathway.
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Capture and analysis of specific ubiquitinylated protein conjugates of interest from particular cell/tissue lysates.
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Purification/pull down of ubiquitinylated proteins from cell free in vitro assays; for example to isolate Ub-p53 species from ubiquitinylation assays mediated by HeLa S100 lysate (Prod. No. BML-SW8750).
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Release of free proteins in their active/native form by cleavage of ubiquitin/ubiquitin chains from the UBI-QAPTURE-Q® matrix using deubiquitinylating enzymes such as USP2 (Prod. No. BML-UW9850).
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Release of ubiquitinylated proteins in their active/native form by elution from the UBI-QAPTURE-Q® matrix using, for example, high salt buffer.
Note: Protocol provided covers applications 1-3. Assay set-up can be readily modified for alternative applications by inclusion, omission or substitution of specific components. Kit is not suitable for use with purified proteins. |
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| Quantity: | Provides sufficient material for 20 x 20μl reactions. |
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| Use/Stability: | Upon receipt, the control lysate (Prod. No. BML-UW0130) should be stored at -80°C. All other components should be stored at 4°C. |
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| Shipping: | Shipped on Dry Ice |
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| Long Term Storage: | -80°C |
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| Contents: | UBI-QAPTURE-Q® matrix (Prod. No. BML-UW0125) 0.5 ml settled resin provided; 50% suspension.
Control ubiquitinylated-protein lysate (Ub-lysate) (Prod. No. BML-UW0130) 500 µg (100µl), 5mg/ml in 50mM HEPES, pH 7.6, 1mM DTT.
Mono- and polyubiquitinylated conjugates monoclonal antibody (UBCJ2) (HRP conjugate) (Ubiquitin-conjugates specific HRP-linked antibody solution) (Prod. No. ENZ-ABS840HRP) 25µl. |
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| Regulatory Status: | RUO - Research Use Only |
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Product Literature References
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Ubiquitination of the Cytoplasmic Domain of Influenza A Virus M2 Protein Is Crucial for Production of Infectious Virus Particles: W.C. Su, et al.; J. Virol.
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Aging Triggers Cytoplasmic Depletion and Nuclear Translocation of the E3 Ligase Mahogunin: A Function for Ubiquitin in Neuronal Survival: S. Benvegnu, et al.; Mol. Cell
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Antiglioma effects of N6-isopentenyladenosine, an endogenous isoprenoid end product, through the downregulation of epidermal growth factor receptor: E. Ciaglia, et al.; Int. J. Cancer
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miR-100 antagonism triggers apoptosis by inhibiting ubiquitination-mediated p53 degradation: G. Yang, et al.; Oncogene
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Mycobacterium tuberculosis Controls Phagosomal Acidification by Targeting CISH-Mediated Signaling: C.J. Queval, et al.; Cell Rep.
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The CreB deubiquitinating enzyme does not directly target the CreA repressor protein in Aspergillus nidulans: M.A. Alam, et al.; Curr. Genet.
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The Pepper RING-Type E3 Ligase CaAIRF1 Regulates ABA and Drought Signaling via CaADIP1 Protein Phosphatase Degradation: C.W. Lim, et al.; Plant Physiol.
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The ubiquitin-conjugating enzyme CDC34 is essential for cytokinesis in contrast to putative subunits of a SCF complex in Trypanosoma brucei: F. Rojas, et al.; PLoS Negl. Trop. Dis.
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E11/podoplanin protein stabilization through inhibition of the proteasome promotes osteocyte differentiation in murine in vitro models: K.A. Staines, et al.; J. Cell. Physiol.
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The ubiquitination of serotonin transporter in lymphoblasts derived from fluvoxamine-resistant depression patients: A. Mouri, et al.; Neurosci. Lett.
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PI3K/AKT signaling inhibits NOTCH1 lysosome‐mediated degradation: N. Platonova, et al.; Genes Chromosomes Cancer
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SKP2 cooperates with N-Ras or AKT to induce liver tumor development in mice: S. Delogu, et al.; Oncotarget
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TRPM8 channel as a novel molecular target in androgen-regulated prostate cancer cells: S. Asuthkar, et al.; Oncotarget
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A Heart That Beats for 500 Years: Age-Related Changes in Cardiac Proteasome Activity, Oxidative Protein Damage and Expression of Heat Shock Proteins, Inflammatory Factors, and Mitochondrial Complexes in Arctica islandica, the Longest-Living Noncolonial Animal: D. Sosnowska, et al.; J Gerontol A Biol Sci Med Sci
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Chloroquine reduces osteoclastogenesis in murine osteoporosis by preventing TRAF3 degradation: Y. Xiu, et al.; J. Clin. Invest.
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HIV-1 Nef down-modulates C-C and C-X-C chemokine receptors via ubiquitin and ubiquitin-independent mechanism: P. Chandrasekaran, et al.; PLoS One
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Nuclear translocation of Hand-1 acts as a molecular switch to regulate vascular radiosensitivity in medulloblastoma tumors: the protein uPAR is a cytoplasmic sequestration factor for Hand-1: S. Asuthkar, et al.; Mol. Cancer Ther.
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Activin receptor-like kinase5 inhibition suppresses mouse melanoma by ubiquitin degradation of Smad4, thereby derepressing eomesodermin in cytotoxic T lymphocytes: J.H. Yoon, et al.; EMBO Mol. Med.
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Deletion of the Type II TGF-β receptor gene in articular chondrocytes leads to a progressive OA-like phenotype in mice: J. Shen, et al.; Arthritis Rheum.
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The Giardia cell cycle progresses independently of the anaphase-promoting complex: S. Gourguechon, et al.; J. Cell Sci.
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Ubiquitin E3 Ligase Itch Negatively Regulates Osteoclast Formation by Promoting Deubiquitination of Tumor Necrosis Factor (TNF) Receptor-associated Factor 6: H. Zhang, et al.; J. Biol. Chem.
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Chaperones, but not oxidized proteins, are ubiquitinated after oxidative stress: M. Kästle, et al.; Free Radic. Biol. Med.
53, 1468 (2012),
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MAGE-D1 regulates expression of depression-like behavior through serotonin transport ubiquitylation: A. Mouri, et al.; J. Neurosci
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Proteomic analysis of ubiquitination-associated proteins in a cisplatin-resistant human lung adenocarcinoma cell line: X. Qin, et al.; Int. J. Mol. Med.
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Ubiquitin-mediated control of plant hormone signaling: D.R. Kelley & M. Estelle; Plant Physiol.
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A tetrapyrrole-regulated ubiquitin ligase controls algal nuclear DNA replication: Y. Kobayashi, et al.; Nat. Cell. Biol.
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BMP2, but not BMP4, is crucial for chondrocyte proliferation and maturation during endochondral bone development: B. Shu, et al.; J. Cell Sci.
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Neuronal precursor cell-expressed developmentally down-regulated 4-1 (NEDD4-1) controls the sorting of newly synthesized Cav1.2 calcium channels: J. Rougier, et al.; J. Biol. Chem.
286, 8829 (2011),
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Proteins bearing oxidation-induced carbonyl groups are not preferentially ubiquitinated: M. Kästle & T. Grune; Biochimie
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The role of the ubiquitin proteasome system in Alzheimer's disease: B.M. Riederer, et al.; Exp. Biol. Med.
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Cross-talk between remodeling and de novo pathways maintains phospholipid balance through ubiquitination: P.L. Butler & R.K. Mallampalli; J. Biol. Chem.
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Smad3 prevents beta-catenin degradation and facilitates beta-catenin nuclear translocation in chondrocytes: M. Zhang, et al.; J. Biol. Chem.
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BAX/BAK-independent mitoptosis during cell death induced by proteasome inhibition: E. Lomonosova, et al.; Mol. Cancer Res.
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Homologous recombination but not nucleotide excision repair plays a pivotal role in tolerance of DNA-protein cross-links in mammalian cells: T. Nakano, et al.; J. Biol. Chem.
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