Validated using a knockout cell lineRecombinantRabMAb

Recombinant Anti-OTUB1 antibody [EPR13028(B)] (ab175200)

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Western blot - Anti-OTUB1 antibody [EPR13028(B)] (ab175200)
  • Western blot - Anti-OTUB1 antibody [EPR13028(B)] (ab175200)
  • Immunoprecipitation - Anti-OTUB1 antibody [EPR13028(B)] (ab175200)
  • Western blot - Anti-OTUB1 antibody [EPR13028(B)] (ab175200)
  • Immunoprecipitation - Anti-OTUB1 antibody [EPR13028(B)] (ab175200)
  • Western blot - Anti-OTUB1 antibody [EPR13028(B)] (ab175200)
  • Anti-OTUB1 antibody [EPR13028(B)] (ab175200)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR13028(B)] to OTUB1
  • Suitable for: WB, IP
  • Knockout validated
  • Reacts with: Mouse, Rat, Human

Conjugates logo Related conjugates and formulations

Carrier Free

Overview

  • Product name

    Anti-OTUB1 antibody [EPR13028(B)]
    See all OTUB1 primary antibodies

  • Description

    Rabbit monoclonal [EPR13028(B)] to OTUB1

  • Host species

    Rabbit

  • Tested applications

    Suitable for: WB, IPmore details
    Unsuitable for: Flow Cyt,ICC/IF or IHC-P

  • Species reactivity

    Reacts with: Mouse, Rat, Human

  • Immunogen

    Synthetic peptide within Human OTUB1 aa 200 to the C-terminus (Cysteine residue). The exact sequence is proprietary.
    Database link: Q96FW1

  • Positive control

    • WB: Wild-type HAP1, HeLa, MCF7, HepG2, HEK-293T, and HEK-293 cell lysates. Rat and mouse heart tissue lysates. IP: HeLa cell lysate.

  • General notes

    This product is a recombinant monoclonal antibody, which offers several advantages including:

    • - High batch-to-batch consistency and reproducibility
    • - Improved sensitivity and specificity
    • - Long-term security of supply
    • - Animal-free production
    For more information see here.

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

Properties

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab175200 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
WB (2)
1/1000 - 1/10000. Predicted molecular weight: 31 kDa.
IP (1)
1/10 - 1/100.
Notes
WB
1/1000 - 1/10000. Predicted molecular weight: 31 kDa.
IP
1/10 - 1/100.
Application notes
Is unsuitable for Flow Cyt,ICC/IF or IHC-P.

Target

  • Function

    Hydrolase that can remove conjugated ubiquitin from proteins and plays an important regulatory role at the level of protein turnover by preventing degradation. Regulator of T-cell anergy, a phenomenon that occurs when T-cells are rendered unresponsive to antigen rechallenge and no longer respond to their cognate antigen. Acts via its interaction with RNF128/GRAIL, a crucial inductor of CD4 T-cell anergy. Isoform 1 destabilizes RNF128, leading to prevent anergy. In contrast, isoform 2 stabilizes RNF128 and promotes anergy. Surprisingly, it regulates RNF128-mediated ubiquitination, but does not deubiquitinate polyubiquitinated RNF128. Deubiquitinates estrogen receptor alpha (ESR1). Mediates deubiquitination of 'Lys-48'-linked polyubiquitin chains, but not 'Lys-63'-linked polyubiquitin chains. Not able to cleave di-ubiquitin. Also capable of removing NEDD8 from NEDD8 conjugates, but with a nuch lower preference compared to 'Lys-48'-linked ubiquitin.

  • Tissue specificity

    Isoform 1 is ubiquitous. Isoform 2 is expressed only in lymphoid tissues such as tonsils, lymph nodes and spleen, as well as peripheral blood mononuclear cells.

  • Sequence similarities

    Belongs to the peptidase C65 family.
    Contains 1 OTU domain.

  • Domain

    In addition to ubiquitin-binding at the Cys-91 active site, a proximal ubiquitin-binding site is also present at Cys-23 Occupancy of the active site is needed to enable tight binding to the second site. Distinct binding sites for the ubiquitins may allow to discriminate among different isopeptide linkages (i.e. 'Lys-48'-, 'Lys-63'-linked polyubiquitin) in polyubiquitin substrates and achieve linkage-specific deubiquitination.

  • Cellular localization

    Cytoplasm.
  • Information by UniProt

  • Database links

    see all

  • Alternative names

    • Deubiquitinating enzyme OTUB1 antibody
    • hOTU1 antibody
    • HSPC263 antibody
    • OTB1 antibody
    • OTU domain containing ubiquitin aldehyde binding protein 1 antibody
    • OTU domain, ubiquitin aldehyde binding 1 antibody
    • OTU domain-containing Ubal-binding protein 1 antibody
    • OTU domain-containing ubiquitin aldehyde-binding protein 1 antibody
    • OTU-domain Ubal-binding 1 antibody
    • OTU1 antibody
    • Otub1 antibody
    • OTUB1_HUMAN antibody
    • Otubain 1 antibody
    • Otubain-1 antibody
    • Ubiquitin specific processing protease OTUB1 antibody
    • Ubiquitin thioesterase OTUB1 antibody
    • ubiquitin-specific protease otubain 1 antibody
    • Ubiquitin-specific-processing protease OTUB1 antibody
    see all

Images

  • Western blot - Anti-OTUB1 antibody [EPR13028(B)] (ab175200)
    Western blot - Anti-OTUB1 antibody [EPR13028(B)] (ab175200)
    All lanes : Anti-OTUB1 antibody [EPR13028(B)] (ab175200) at 1/1000 dilution

    Lane 1 : Wild-type HEK-293T cell lysate
    Lane 2 : OTUB1 knockout HEK-293T cell lysate

    Lysates/proteins at 20 µg per lane.

    Performed under reducing conditions.

    Predicted band size: 31 kDa
    Observed band size: 31 kDa



    Lanes 1-2: Merged signal (red and green). Green - ab175200 observed at 31 kDa. Red - loading control ab8245 observed at 37 kDa.

     ab175200 Anti-OTUB1 antibody [EPR13028(B)] was shown to specifically react with OTUB1 in wild-type HEK-293T cells. Loss of signal was observed when knockout cell line ab266551 (knockout cell lysate ab257569) was used. Wild-type and OTUB1 knockout samples were subjected to SDS-PAGE. ab175200 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°CC at 1 in 1000 Dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

     

  • Western blot - Anti-OTUB1 antibody [EPR13028(B)] (ab175200)
    Western blot - Anti-OTUB1 antibody [EPR13028(B)] (ab175200)
    All lanes : Anti-OTUB1 antibody [EPR13028(B)] (ab175200) at 1/1000 dilution

    Lane 1 : Wild-type HAP1 whole cell lysate
    Lane 2 : OTUB1 knockout HAP1 whole cell lysate
    Lane 3 : HeLa whole cell lysate
    Lane 4 : HEK-293 whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Predicted band size: 31 kDa



    Lanes 1 - 4: Merged signal (red and green). Green - ab175200 observed at 35 kDa. Red - loading control, ab18058, observed at 130 kDa.

    ab175200 was shown to specifically react with OTUB1 in wild type cells as signal was lost in OTUB1 knockout cells. Wild-type and OTUB1 knockout samples were subjected to SDS-PAGE. ab175200 and ab18058 (Mouse anti-Vinculin loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.

  • Immunoprecipitation - Anti-OTUB1 antibody [EPR13028(B)] (ab175200)
    Immunoprecipitation - Anti-OTUB1 antibody [EPR13028(B)] (ab175200)

    ab175200 (purified) at 1:50 dilution (2ug) immunoprecipitating OTUB1 in HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate.
    Lane 1 (input): HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate 10ug
    Lane 2 (+): ab175200 & HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate
    Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab175200 in HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate
    For western blotting, VeriBlot for IP Detection Reagent (HRP) (ab131366) was used for detection at 1:1000 dilution.
    Blocking and diluting buffer: 5% NFDM/TBST.

  • Western blot - Anti-OTUB1 antibody [EPR13028(B)] (ab175200)
    Western blot - Anti-OTUB1 antibody [EPR13028(B)] (ab175200)
    All lanes : Anti-OTUB1 antibody [EPR13028(B)] (ab175200) at 1/1000 dilution (purified)

    Lane 1 : HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysates
    Lane 2 : Mouse heart whole tissue lysates
    Lane 3 : Rat heart whole tissue lysates

    Lysates/proteins at 15 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

    Predicted band size: 31 kDa
    Observed band size: 31 kDa



    Blocking and diluting buffer: 5% NFDM/TBST

  • Immunoprecipitation - Anti-OTUB1 antibody [EPR13028(B)] (ab175200)
    Immunoprecipitation - Anti-OTUB1 antibody [EPR13028(B)] (ab175200)

    Western blot analysis on immunoprecipitation pellet from MCF7 cell lysate labeling OTUB1 with unpurified ab175200 at 1/10 dilution.

  • Western blot - Anti-OTUB1 antibody [EPR13028(B)] (ab175200)
    Western blot - Anti-OTUB1 antibody [EPR13028(B)] (ab175200)
    All lanes : Anti-OTUB1 antibody [EPR13028(B)] (ab175200) at 1/1000 dilution (unpurified)

    Lane 1 : HeLa cell lysate
    Lane 2 : HepG2 cell lysate
    Lane 3 : 293T cell lysate
    Lane 4 : MCF7 cell lysate

    Lysates/proteins at 10 µg per lane.

    Predicted band size: 31 kDa

  • Anti-OTUB1 antibody [EPR13028(B)] (ab175200)
    Anti-OTUB1 antibody [EPR13028(B)] (ab175200)

References (6)

Publishing research using ab175200? Please let us know so that we can cite the reference in this datasheet.

ab175200 has been referenced in 6 publications.

  • Yang L  et al. Broad Spectrum Deubiquitinase Inhibition Induces Both Apoptosis and Ferroptosis in Cancer Cells. Front Oncol 10:949 (2020). PubMed: 32596160
  • Xu Y  et al. Anti-bacterial and anti-viral nanchangmycin displays anti-myeloma activity by targeting Otub1 and c-Maf. Cell Death Dis 11:818 (2020). PubMed: 32999280
  • Liang C  et al. LncRNA GAS6-AS2 facilitates tumor growth and metastasis of hepatocellular carcinoma by activating the PI3K/AKT/FoxO3a signaling pathway. Int J Clin Exp Pathol 12:4011-4023 (2019). PubMed: 31933797
  • Zhou X  et al. The deubiquitinase Otub1 controls the activation of CD8+ T cells and NK cells by regulating IL-15-mediated priming. Nat Immunol 20:879-889 (2019). PubMed: 31182807
  • Liu T  et al. The Deubiquitylase OTUB1 Mediates Ferroptosis via Stabilization of SLC7A11. Cancer Res 79:1913-1924 (2019). PubMed: 30709928
  • Pasupala N  et al. OTUB1 non-catalytically stabilizes the E2 ubiquitin-conjugating enzyme UBE2E1 by preventing its autoubiquitination. J Biol Chem 293:18285-18295 (2018). PubMed: 30282802

Customer reviews and Q&As

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Immunoprecipitation abreview for Anti-OTUB1 [EPR13028(B)] antibody

 Excellent
Abreviews
abreview image
Application
Immunoprecipitation
Immuno-precipitation step
Protein A
Sample
Human Cell lysate - whole cell (HEK293 cells)
Specification
HEK293 cells
Total protein in input
20 µg
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Submitted Nov 12 2013

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