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  1. Link

    products/primary-antibodies/otub1-antibody-ab101471.pdf

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Epigenetics and Nuclear Signaling DNA / RNA DNA Damage & Repair Homologous Recomb.
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Anti-OTUB1 antibody (ab101471)

  • Datasheet
  • SDS
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Western blot - Anti-OTUB1 antibody (ab101471)
  • Western blot - Anti-OTUB1 antibody (ab101471)
  • Western blot - Anti-OTUB1 antibody (ab101471)
  • Immunocytochemistry/ Immunofluorescence - Anti-OTUB1 antibody (ab101471)

Key features and details

  • Rabbit polyclonal to OTUB1
  • Suitable for: WB, ICC/IF
  • Reacts with: Mouse, Human
  • Isotype: IgG

You may also be interested in

Primary
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Anti-TNFAIP3 antibody [59A426] (ab13597)
Secondary
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Goat Anti-Rabbit IgG H&L (HRP) (ab205718)
Knockout
Product image
Human OTUB1 knockout HEK-293T cell line (ab266551)

View more associated products

Overview

  • Product name

    Anti-OTUB1 antibody
    See all OTUB1 primary antibodies
  • Description

    Rabbit polyclonal to OTUB1
  • Host species

    Rabbit
  • Tested applications

    Suitable for: WB, ICC/IFmore details
  • Species reactivity

    Reacts with: Mouse, Human
    Predicted to work with: Rat, Dog, Pig
  • Immunogen

    Synthetic peptide corresponding to Human OTUB1 aa 200 to the C-terminus conjugated to keyhole limpet haemocyanin.
    (Peptide available as ab113298)

  • Positive control

    • WB: HEK-293T, HAP1 and HeLa cell lysate; Mouse brain tissue lysate. ICC/IF: HeLa cells.
  • General notes

    The Life Science industry has been in the grips of a reproducibility crisis for a number of years. Abcam is leading the way in addressing this with our range of recombinant monoclonal antibodies and knockout edited cell lines for gold-standard validation. Please check that this product meets your needs before purchasing.

    If you have any questions, special requirements or concerns, please send us an inquiry and/or contact our Support team ahead of purchase. Recommended alternatives for this product can be found below, along with publications, customer reviews and Q&As

Properties

  • Form

    Liquid
  • Storage instructions

    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C or -80°C. Avoid freeze / thaw cycle.
  • Storage buffer

    pH: 7.40
    Preservative: 0.02% Sodium azide
    Constituent: PBS

    Batches of this product that have a concentration < 1mg/ml may have BSA added as a stabilising agent. If you would like information about the formulation of a specific lot, please contact our scientific support team who will be happy to help.
  • Concentration information loading...
  • Purity

    Immunogen affinity purified
  • Clonality

    Polyclonal
  • Isotype

    IgG
  • Research areas

    • Epigenetics and Nuclear Signaling
    • DNA / RNA
    • DNA Damage & Repair
    • Homologous Recomb.
    • Cell Biology
    • Proteolysis / Ubiquitin
    • Proteasome / Ubiquitin
    • Deubiquitination
    • Cell Biology
    • Proteolysis / Ubiquitin
    • Proteolytic enzymes
    • Other proteases
    • Epigenetics and Nuclear Signaling
    • Ubiquitin & Ubiquitin Like Modifiers
    • Deubiquitination

Associated products

  • Compatible Secondaries

    • Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077)
    • Goat Anti-Rabbit IgG H&L (HRP) (ab205718)
  • Isotype control

    • Rabbit IgG, polyclonal - Isotype Control (ChIP Grade) (ab171870)
  • KO cell lines

    • Human OTUB1 knockout HEK-293T cell line (ab266551)
  • KO cell lysates

    • Human OTUB1 knockout HEK-293T cell lysate (ab257569)
  • Recombinant Protein

    • Recombinant human OTUB1 protein (ab157086)

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab101471 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 (1)
Use a concentration of 1 µg/ml. Detects a band of approximately 31 kDa (predicted molecular weight: 31 kDa).
ICC/IF
Use a concentration of 5 µg/ml.
Notes
WB
Use a concentration of 1 µg/ml. Detects a band of approximately 31 kDa (predicted molecular weight: 31 kDa).
ICC/IF
Use a concentration of 5 µg/ml.

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.
  • Target information above from: UniProt accession Q96FW1 The UniProt Consortium
    The Universal Protein Resource (UniProt) in 2010
    Nucleic Acids Res. 38:D142-D148 (2010) .

    Information by UniProt
  • Database links

    • Entrez Gene: 55611 Human
    • Entrez Gene: 107260 Mouse
    • Entrez Gene: 100302092 Pig
    • Entrez Gene: 293705 Rat
    • Omim: 608337 Human
    • SwissProt: Q96FW1 Human
    • SwissProt: Q7TQI3 Mouse
    • SwissProt: B2RYG6 Rat
    • Unigene: 473788 Human
    • Unigene: 203921 Mouse
    • Unigene: 9218 Rat
    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 (ab101471)
    Western blot - Anti-OTUB1 antibody (ab101471)
    All lanes : Anti-OTUB1 antibody (ab101471) at 1/10000 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: 130 kDa why is the actual band size different from the predicted?



    Lanes 1-2: Merged signal (red and green). Green - ab101471 observed at 130 kDa. Red - loading control ab8245 observed at 37 kDa.

     ab101471 Anti-OTUB1 antibody 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. ab101471 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 10000 ug/ml 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 (ab101471)
    Western blot - Anti-OTUB1 antibody (ab101471)
    All lanes : Anti-OTUB1 antibody (ab101471) at 1 µg/ml

    Lane 1 : Wild-type HAP1 whole cell lysate
    Lane 2 : OTUB1 knockout HAP1 whole cell lysate
    Lane 3 : HeLa whole cell lysate
    Lane 4 : Hek293 whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Predicted band size: 31 kDa



    Lanes 1 - 4: Merged signal (red and green). Green - ab101471 observed at 31 kDa. Red - loading control, ab18058, observed at 130 kDa.

    ab101471 was shown to recognize OTUB1 in wild-type HAP1 cells as signal was lost at the expected MW in OTUB1 knockout cells. Additional cross-reactive bands were observed in the wild-type and knockout cells. Wild-type and OTUB1 knockout samples were subjected to SDS-PAGE. ab101471 and ab18058 (Mouse anti-Vinculin loading control) were incubated overnight at 4°C at 1µg/mL 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.

  • Western blot - Anti-OTUB1 antibody (ab101471)
    Western blot - Anti-OTUB1 antibody (ab101471)
    All lanes : Anti-OTUB1 antibody (ab101471) at 1 µg/ml

    Lane 1 : Brain (Mouse) Tissue Lysate
    Lane 2 : Human brain tissue lysate - total protein (ab29466)

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 dilution

    Developed using the ECL technique.

    Performed under reducing conditions.

    Predicted band size: 31 kDa
    Observed band size: 31 kDa
    Additional bands at: 62 kDa. We are unsure as to the identity of these extra bands.


    Exposure time: 12 minutes


    Secondary antibody - goat anti-rabbit HRP (ab97080)

  • Immunocytochemistry/ Immunofluorescence - Anti-OTUB1 antibody (ab101471)
    Immunocytochemistry/ Immunofluorescence - Anti-OTUB1 antibody (ab101471)

    ICC/IF image of ab101471 stained HeLa cells. The cells were 4% PFA fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab101471, 5µg/ml) overnight at +4°C. The secondary antibody (green) was ab96899, goat anti-rabbit DyLight® 488 (IgG H+L) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM. This antibody also gave a positive result in 4% PFA fixed (10 min) Hek293, HepG2 and MCF7 cells at 5µg/ml, and in 100% methanol fixed (5 min)HeLa, HepG2 and MCF7 cells at 5µg/ml.

Protocols

  • Western blot protocols
  • Immunocytochemistry & immunofluorescence protocols

Click here to view the general protocols

Datasheets and documents

  • SDS download

  • Datasheet download

    Download

References (0)

Publishing research using ab101471? Please let us know so that we can cite the reference in this datasheet.

ab101471 has not yet been referenced specifically in any publications.

Customer reviews and Q&As

Show All Reviews Q&A
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1-3 of 3 Abreviews or Q&A

Flow Cytometry abreview for Anti-OTUB1 antibody

Excellent
Abreviews
Abreviews
abreview image
Application
Flow Cytometry
Sample
Mouse Cell (spleen CD4 T cells)
Permeabilization
Yes - Foxp3 Fixation/Permeabilization
Gating Strategy
viable cells
Specification
spleen CD4 T cells
Fixation
Foxp3 Fixation/Permeabilization
Read More
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

Dr. Cinthia Stempin

Verified customer

Submitted Aug 03 2018

Immunoprecipitation abreview for Anti-OTUB1 antibody

Poor
Abreviews
Abreviews
abreview image
Application
Immunoprecipitation
Sample
Human Cell lysate - whole cell (HEK293 cells)
Total protein in input
20 µg
Immuno-precipitation step
Protein A
Specification
HEK293 cells
Read More
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

Abcam user community

Verified customer

Submitted Dec 02 2013

Western blot abreview for Anti-OTUB1 antibody

Excellent
Abreviews
Abreviews
abreview image
Application
Western blot
Loading amount
20 µg
Gel Running Conditions
Reduced Denaturing
Sample
Human Cell lysate - whole cell (HEK293 cells)
Specification
HEK293 cells
Treatment
untreated or HA-OTUB1 transfected for 48 hours
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 21°C
Read More
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

Abcam user community

Verified customer

Submitted Nov 12 2013

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