Overview

  • Product nameAnti-YB1 antibody [EP2708Y]
    See all YB1 primary antibodies
  • Description
    Rabbit monoclonal [EP2708Y] to YB1
  • Tested applicationsSuitable for: ICC/IF, WB, IP, IHC-P, Flow Cytmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide (the amino acid sequence is considered to be commercially sensitive) within Human YB1 aa 250 to the C-terminus (C terminal). The exact sequence is proprietary.
    (Peptide available as ab175051)

  • Positive control
    • WB: HeLa, SW480, A549, C6 PC-12, NIH/3T3, Raw264.7 and MCF7 cell lysates. IHC-P: Human kidney, human cervical carcinoma, mouse liver and rat stomach tissues. ICC/IF: HeLa cells. Flow Cyt: HeLa cells. IP: HEK293, HeLa and MCF-7 whole cell lysate.
  • General notes

    This product is a recombinant rabbit monoclonal antibody.

     

    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.

    Produced using Abcam’s RabMAb® technology. RabMAb® technology is covered by the following U.S. Patents, No. 5,675,063 and/or 7,429,487.

    Alternative versions available:

    Anti-YB1 antibody (HRP) [EP2708Y] (ab204014)

Properties

Applications

Our Abpromise guarantee covers the use of ab76149 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
ICC/IF 1/100.

For unpurified use at 5µg/ml.

WB 1/1000. Predicted molecular weight: 36 kDa.Can be blocked with YB1 peptide (ab175051).

For unpurified use at 1/10000 - 1/20000.

IP 1/30.
IHC-P 1/50 - 1/250. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

See protocols (link: http://www.abcam.com/protocols/ihc-antigen-retrieval-protocol).

Flow Cyt 1/50 - 1/100.

ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

Target

  • FunctionMediates pre-mRNA alternative splicing regulation. Binds to splice sites in pre-mRNA and regulates splice site selection. Binds and stabilizes cytoplasmic mRNA. Contributes to the regulation of translation by modulating the interaction between the mRNA and eukaryotic initiation factors (By similarity). Regulates the transcription of numerous genes. Its transcriptional activity on the multidrug resistance gene MDR1 is enhanced in presence of the APEX1 acetylated form at 'Lys-6' and 'Lys-7'. Binds to promoters that contain a Y-box (5'-CTGATTGGCCAA-3'), such as MDR1 and HLA class II genes. Promotes separation of DNA strands that contain mismatches or are modified by cisplatin. Has endonucleolytic activity and can introduce nicks or breaks into double-stranded DNA (in vitro). May play a role in DNA repair. Component of the CRD-mediated complex that promotes MYC mRNA stability.
    The secreted form acts as an extracellular mitogen and stimulates cell migration and proliferation.
  • Sequence similaritiesContains 1 CSD (cold-shock) domain.
  • Post-translational
    modifications
    Ubiquitinated by RBBP6; leading to a decrease of YBX1 transcativational ability.
    In the absence of phosphorylation the protein is retained in the cytoplasm.
    Cleaved by a 20S proteasomal protease in response to agents that damage DNA. Cleavage takes place in the absence of ubiquitination and ATP. The resulting N-terminal fragment accumulates in the nucleus.
  • Cellular localizationCytoplasm. Nucleus. Cytoplasmic granule. Secreted. Localized in cytoplasmic mRNP granules containing untranslated mRNAs. Shuttles between nucleus and cytoplasm. Predominantly cytoplasmic in proliferating cells. Cytotoxic stress and DNA damage enhance translocation to the nucleus. Localized with DDX1, MBNL1 and TIAL1 in stress granules upon stress. Secreted by mesangial and monocytic cells after inflammatory challenges. Translocates from the cytoplasm to the nucleus after and colocalizes with APEX1 in nuclear speckles after genotoxic stress.
  • Information by UniProt
  • Database links
  • Alternative names
    • BP 8 antibody
    • CBF-A antibody
    • CCAAT binding transcription factor I subunit A antibody
    • CCAAT-binding transcription factor I subunit A antibody
    • CSDA2 antibody
    • CSDB antibody
    • DBPB antibody
    • DNA binding protein B antibody
    • DNA-binding protein B antibody
    • EFI-A antibody
    • Enhancer factor I subunit A antibody
    • MDR NF1 antibody
    • MGC104858 antibody
    • MGC110976 antibody
    • MGC117250 antibody
    • NSEP 1 antibody
    • NSEP1 antibody
    • Nuclease sensitive element binding protein 1 antibody
    • Nuclease-sensitive element-binding protein 1 antibody
    • p50 antibody
    • Q15905 antibody
    • Y-box binding protein 1 antibody
    • Y-box transcription factor antibody
    • Y-box-binding protein 1 antibody
    • YB 1 antibody
    • YB-1 antibody
    • YBOX1_HUMAN antibody
    • YBX 1 antibody
    • ybx1 antibody
    see all

Anti-YB1 antibody [EP2708Y] images

  • All lanes : Anti-YB1 antibody [EP2708Y] (ab76149) at 1/1000 dilution (purified)

    Lane 1 : NIH/3T3 whole cell lysate
    Lane 2 : Raw264.7 whole cell lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/50000 dilution

    Predicted band size : 36 kDa
    Observed band size : 50 kDa (why is the actual band size different from the predicted?)

    Blocking and dilution buffer: 5% NFDM/TBST.

  • All lanes : Anti-YB1 antibody [EP2708Y] (ab76149) at 1/10000 dilution (purified)

    Lane 1 : C6 whole cell lysate
    Lane 2 : PC-12 whole cell lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/50000 dilution

    Predicted band size : 36 kDa
    Observed band size : 50 kDa (why is the actual band size different from the predicted?)

    Blocking and dilution buffer: 5% NFDM/TBST.

  • All lanes : Anti-YB1 antibody [EP2708Y] (ab76149) at 1/1000 dilution (purified)

    Lane 1 : HeLa whole cell lysate
    Lane 2 : SW480 whole cell lysate
    Lane 3 : A549 whole cell lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/50000 dilution

    Predicted band size : 36 kDa
    Observed band size : 50 kDa (why is the actual band size different from the predicted?)

    Blocking and dilution buffer: 5% NFDM/TBST.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat stomach tissue labelling YB1 with purified ab76149 at 1/50. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of mouse liver tissue labelling YB1 with purified ab76149 at 1/50. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human cervix carcinoma tissue labelling YB1 with purified ab76149 at 1/50. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

  • Immunocytochemistry/Immunofluorescence analysis of HeLa cells labelling YB1 with purified ab76149 at 1/100. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody. DAPI (blue) was used as the nuclear counterstain. ab7291, a mouse anti-tubulin (1/1000) and ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/1000) were also used.

    Control 1: primary antibody (1/100) and secondary antibody, ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/1000).

    Control 2: ab7291 (1/1000) and secondary antibody, ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000).

  • Flow Cytometry analysis of HeLa cells labelling YB1 with purified ab76149 at 1/90 (red). Cells were fixed with 4% paraformaldehyde. A FITC-conjugated goat anti-rabbit IgG (1/500) was used as the secondary antibody. Black - Isotype control, rabbit monoclonal IgG. Blue - Unlabelled control, cells without incubation with primary and secondary antibodies.

  • ab76149 (purified) at 1/30 immunoprecipitating YB1 in MCF-7 whole cell lysate.

    Lane 1 (input): MCF-7 whole cell lysate (10µg)

    Lane 2 (+): ab76149 + MCF-7 whole cell lysate.

    Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab76149 in MCF-7 whole cell lysate.

    For western blotting, a HRP-conjugated anti-rabbit IgG, specific to the non-reduced form of IgG was used as the secondary antibody (1/1500).

    Blocking buffer and concentration: 5% NFDM/TBST.

    Diluting buffer and concentration: 5% NFDM /TBST.

  • ab76149 (purified) at 1/30 immunoprecipitating YB1 in HeLa whole cell lysate.

    Lane 1 (input): HeLa whole cell lysate (10µg)

    Lane 2 (+): ab76149 + HeLa whole cell lysate.

    Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab76149 in HeLa whole cell lysate.

    For western blotting, a HRP-conjugated anti-rabbit IgG, specific to the non-reduced form of IgG was used as the secondary antibody (1/1500).

    Blocking buffer and concentration: 5% NFDM/TBST.

    Diluting buffer and concentration: 5% NFDM /TBST.

  • All lanes : Anti-YB1 antibody [EP2708Y] (ab76149) at 1/200000 dilution (unpurified)

    Lane 1 : HeLa cell lysate
    Lane 2 : SW480 cell lysate
    Lane 3 : A549 cell lysate
    Lane 4 : MCF7 cell lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    HRP-conjugated goat anti-rabbit IgG at 1/1000 dilution

    Predicted band size : 36 kDa
    Observed band size : 50 kDa (why is the actual band size different from the predicted?)
  • ICC/IF image of unpurified ab76149 stained HeLa cells. The cells were 4% formaldehyde 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 (unpurified ab76149, 5µg/ml) overnight at +4°C. The secondary antibody (green) was ab96899, DyLight® 488 goat anti-rabbit 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.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human kidney tissue labelling YB1 with unpurified ab76149 at a dilution of 1/100.

  • Overlay histogram showing HeLa cells stained with unpurified ab76149 (red line). The cells were fixed with 4% paraformaldehyde (10 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (unpurified ab76149, 1/100 dilution) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-rabbit IgG (H+L) (ab96899) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit monoclonal IgG (1µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in HeLa cells fixed with methanol (5 min)/permeabilized with 0.1% PBS-Tween 20 used under the same conditions.

  • YB1 was immunoprecipitated using 0.5mg HEK293 whole cell extract, 10µg of Rabbit monoclonal to YB1 and 50µl of protein G magnetic beads (+). No antibody was added to the control (-).
    The antibody was incubated under agitation with Protein G beads for 10min, HEK293 whole cell extract lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation.
    Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70°C; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with unpurified ab76149.
    Secondary: Mouse monoclonal [SB62a] secondary antibody to rabbit IgG light chain (HRP) (ab99697).
    Band: 46kDa: YB1.

References for Anti-YB1 antibody [EP2708Y] (ab76149)

This product has been referenced in:
  • Liu J  et al. Long noncoding RNA POU6F2-AS2 is associated with oesophageal squamous cell carcinoma. J Biochem 160:195-204 (2016). Read more (PubMed: 27033944) »
  • Imada K  et al. FOXO3a Expression Regulated by ERK Signaling is Inversely Correlated With Y-Box Binding Protein-1 Expression in Prostate Cancer. Prostate N/A:N/A (2016). Read more (PubMed: 27699813) »

See all 6 Publications for this product

Product Wall

Thank you for your enquiry. I can confirm that these products are sold as tissue culture supernatant. Unpurified antibodies, such as those sold as whole antiserum, ascites or tissue culture supernatant generally will not have a concentration stated ...

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Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Human Cell lysate - whole cell (Renal proximal tubular epithelial cells)
Loading amount 10 µg
Specification Renal proximal tubular epithelial cells
Gel Running Conditions Reduced Denaturing (7.5 %)
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 21°C
Username

Dr. Robert Jenkins

Verified customer

Submitted Apr 20 2009

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"