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UBIQAPTURE-Q® kit

Market-leading tool for isolation and detection of ubiquitinylated proteins.
 
BML-UW8995-0001 20 tests 461.00 USD
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  • Efficient tool for the selective isolation of ubiquitinylated proteins
  • High binding affinity matrix, able to bind both monoubiquitinated and polyubiquitinated proteins
  • 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.
BML-UW8995 WB
Western blot analysis of ubiquitin enrichment of lysate derived ubiquitinylated proteins. Ubiquitin enrichment experiment set-up and run as described in “Capture/enrichment of ubiquitinylated proteins”. Ubiquitin-protein conjugates present in starting material, unbound fraction and elution fraction were detected by western blotting as described in “Analysis by western blotting”, using the provided ubiquitin-conjugate specific HRP-linked antibody (Prod. No. BML-UW0150) at a dilution of 1:1000 dilution. Capture of Ub-protein conjugates from Control ubiquitinylated-protein lysate (Prod. No. BML-UW0130). Key: SM = Starting Material, UF = Unbound Fraction and EL = Elution Fraction.
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BML-UW8995 WB

Product Specification

Application Notes:Uses:
  1. 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.
  2. Capture and analysis of specific ubiquitinylated protein conjugates of interest from particular cell/tissue lysates.
  3. 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).
  4. 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).
  5. 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.
 
Quantity:Provides sufficient material for 20 x 20μl reactions.
 
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.
 
Shipping:Shipped on Dry Ice
 
Long Term Storage:-80°C
 
Kit/Set Contains: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.
Ubiquitin-conjugates specific HRP-linked antibody solutions (Prod. No. BML-PW0150) 25µl; HRP-linked antibody for use with ECL western blotting detection reagents.
 

Product Literature References

miR-100 antagonism triggers apoptosis by inhibiting ubiquitination-mediated p53 degradation: G. Yang, et al.; Oncogene 36, 1023 (2017), Application(s): Detection of ubiquitinated protein, Abstract;
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. 231, 1392 (2016), Abstract; Full Text
The ubiquitination of serotonin transporter in lymphoblasts derived from fluvoxamine-resistant depression patients: A. Mouri, et al.; Neurosci. Lett. 617, 22 (2016), Application(s): Detection of ubiquitinylated protein, Abstract;
PI3K/AKT signaling inhibits NOTCH1 lysosome‐mediated degradation: N. Platonova, et al.; Genes Chromosomes Cancer (2015), Abstract;
SKP2 cooperates with N-Ras or AKT to induce liver tumor development in mice: S. Delogu, et al.; Oncotarget 6, 2222 (2015), Abstract; Full Text
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 69, 1448 (2014), Abstract;
Chloroquine reduces osteoclastogenesis in murine osteoporosis by preventing TRAF3 degradation: Y. Xiu, et al.; J. Clin. Invest. 124, 297 (2014), Abstract; Full Text
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 9, e86998 (2014), Abstract; Full Text
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. 5, 1720 (2013), Abstract; Full Text
An immunoaffinity purification method for the proteomic analysis of ubiquitinated protein complexes: P. Schwertman, et al.; Anal. Biochem. 440, 227 (2013), Abstract;
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. 65, 3107 (2013), Abstract;
The Giardia cell cycle progresses independently of the anaphase-promoting complex: S. Gourguechon, et al.; J. Cell Sci. 126, 2246 (2013), Abstract;
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. 288, 22359 (2013), Abstract;
Chaperones, but not oxidized proteins, are ubiquitinated after oxidative stress: M. Kästle, et al.; Free Radic. Biol. Med. 53, 1468 (2012), Abstract;
MAGE-D1 regulates expression of depression-like behavior through serotonin transport ubiquitylation: A. Mouri, et al.; J. Neurosci 32, 4562 (2012), Abstract;
Proteomic analysis of ubiquitination-associated proteins in a cisplatin-resistant human lung adenocarcinoma cell line: X. Qin, et al.; Int. J. Mol. Med. 29, 791 (2012), Abstract;
Ubiquitin-mediated control of plant hormone signaling: D.R. Kelley & M. Estelle; Plant Physiol. 160, 47 (2012), Abstract;
A tetrapyrrole-regulated ubiquitin ligase controls algal nuclear DNA replication: Y. Kobayashi, et al.; Nat. Cell. Biol. 13, 483 (2011), Abstract;
BMP2, but not BMP4, is crucial for chondrocyte proliferation and maturation during endochondral bone development: B. Shu, et al.; J. Cell Sci. 124, 3428 (2011), Abstract;
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), Abstract;
Proteins bearing oxidation-induced carbonyl groups are not preferentially ubiquitinated: M. Kästle, et al.; Biochimie 93, 1076 (2011), Abstract;
The role of the ubiquitin proteasome system in Alzheimer's disease: B.M. Riederer, et al.; Exp. Biol. Med. 236, 268 (2011), Abstract;
Cross-talk between remodeling and de novo pathways maintains phospholipid balance through ubiquitination: P.L. Butler & R.K. Mallampalli; J. Biol. Chem. 285, 6246 (2010), Abstract;
Smad3 prevents beta-catenin degradation and facilitates beta-catenin nuclear translocation in chondrocytes: M. Zhang, et al.; J. Biol. Chem. 285, 8703 (2010), Abstract;
BAX/BAK-independent mitoptosis during cell death induced by proteasome inhibition: E. Lomonosova, et al.; Mol. Cancer Res. 7, 1268 (2009), Abstract;
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. 284, 27065 (2009), Abstract;

General Literature References

BAX/BAK-independent mitoptosis during cell death induced by proteasome inhibition?: E. Lomonosova, et al.; Mol. Cancer Res. 7, 1268 (2009), Abstract;
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. 284, 27065 (2009), Abstract;
Diverse polyubiquitin interaction properties of ubiquitin-associated domains: S. Raasi, et al.; Nat. Struct. Mol. Biol. 12, 708 (2005), Abstract;
Ubiquitin-binding domains: L. Hicke, et al.; Nat. Rev. Mol. Cell. Biol. 6, 610 (2005), Abstract;
Polyubiquitin chains: polymeric protein signals: C.M. Pickart, et al.; Curr. Opin. Chem. Biol. 8, 610 (2004), Abstract;
Mechanisms underlying ubiquitination: C.M. Pickart, et al.; Annu. Rev. Biochem. 70, 503 (2001), Abstract;
Protein regulation by monoubiquitin: L. Hicke, et al.; Nat. Rev. Mol. Cell. Biol. 2, 195 (2001), Abstract;
The ubiquitin-proteasome system and endocytosis: G.J. Strous, et al.; J. Cell. Sci. 112 , 1417 (1999), Abstract;
The ubiquitin system: A. Hershko, et al.; Annu. Rev. Biochem. 67, 425 (1998), Abstract;
Pathways of ubiquitin conjugation: A.L. Haas, et al.; FASEB J. 11, 1257 (1997), Abstract;

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