Validated using a knockout cell line

Anti-Hsp90 beta antibody [H90-10] (ab53497)

Reviews (1)Q&A (2)References (20)
Western blot - Anti-Hsp90 beta antibody [H90-10] (ab53497)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Hsp90 beta antibody [H90-10] (ab53497)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Hsp90 beta antibody [H90-10] (ab53497)
  • Immunocytochemistry/ Immunofluorescence - Anti-Hsp90 beta antibody [H90-10] (ab53497)
  • Western blot - Anti-Hsp90 beta antibody [H90-10] (ab53497)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Hsp90 beta antibody [H90-10] (ab53497)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Hsp90 beta antibody [H90-10] (ab53497)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Hsp90 beta antibody [H90-10] (ab53497)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Hsp90 beta antibody [H90-10] (ab53497)
  • Western blot - Anti-Hsp90 beta antibody [H90-10] (ab53497)
  • Flow Cytometry - Anti-Hsp90 beta antibody [H90-10] (ab53497)

Key features and details

  • Mouse monoclonal [H90-10] to Hsp90 beta
  • Suitable for: Flow Cyt, IHC-P, ICC/IF, WB
  • Knockout validated
  • Reacts with: Mouse, Human
  • Isotype: IgG2a

Overview

  • Product name

    Anti-Hsp90 beta antibody [H90-10]
    See all Hsp90 beta primary antibodies

  • Description

    Mouse monoclonal [H90-10] to Hsp90 beta

  • Host species

    Mouse

  • Specificity

    Detects 90kDa. Detects HSP90 beta in all reactive species except in Chicken, where it detects both alpha and beta isoforms.

  • Tested applications

    Suitable for: Flow Cyt, IHC-P, ICC/IF, WBmore details

  • Species reactivity

    Reacts with: Mouse, Human

  • Immunogen

    Recombinant full length protein corresponding to Human Hsp90 beta.
    Database link: P08238

  • Positive control

    • WB: HEK-293T, Saos-2 and HL-60 cell lysates; Recombinant human Hsp90 beta protein. IHC-P: Human placenta, backskin and colon carcinoma tissue; mouse colon tissue. ICC/IF: HeLa cells. Flow Cyt: 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. Upon delivery aliquot and store at -20°C. Avoid freeze / thaw cycles.

  • Storage buffer

    pH: 7.2
    Preservative: 0.09% Sodium azide
    Constituents: 50% Glycerol (glycerin, glycerine), PBS
  • Concentration information loading...

  • Purity

    Protein G purified

  • Clonality

    Monoclonal

  • Clone number

    H90-10

  • Isotype

    IgG2a

  • Research areas

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab53497 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
Flow Cyt
Use 0.5µg for 106 cells.

ab170191 - Mouse monoclonal IgG2a, is suitable for use as an isotype control with this antibody.

 
IHC-P
1/100.
ICC/IF
Use at an assay dependent concentration.
WB (1)
1/2500. Predicted molecular weight: 83 kDa.
Notes
Flow Cyt
Use 0.5µg for 106 cells.

ab170191 - Mouse monoclonal IgG2a, is suitable for use as an isotype control with this antibody.

 
IHC-P
1/100.
ICC/IF
Use at an assay dependent concentration.
WB
1/2500. Predicted molecular weight: 83 kDa.

Target

  • Function

    Molecular chaperone that promotes the maturation, structural maintenance and proper regulation of specific target proteins involved for instance in cell cycle control and signal transduction. Undergoes a functional cycle that is linked to its ATPase activity. This cycle probably induces conformational changes in the client proteins, thereby causing their activation. Interacts dynamically with various co-chaperones that modulate its substrate recognition, ATPase cycle and chaperone function.

  • Sequence similarities

    Belongs to the heat shock protein 90 family.

  • Domain

    The TPR repeat-binding motif mediates interaction with TPR repeat-containing proteins.

  • Post-translational
    modifications

    Ubiquitinated in the presence of STUB1-UBE2D1 complex (in vitro).
    ISGylated.
    S-nitrosylated; negatively regulates the ATPase activity.

  • Cellular localization

    Cytoplasm. Melanosome. Identified by mass spectrometry in melanosome fractions from stage I to stage IV.
  • Information by UniProt

  • Database links

    • Alternative names

      • 90 kda heat shock protein beta HSP90 beta antibody
      • D6S182 antibody
      • FLJ26984 antibody
      • Heat shock 84 kDa antibody
      • Heat shock 90kD protein 1, beta antibody
      • Heat shock 90kDa protein 1 beta antibody
      • Heat shock protein 90 alpha family class B member 1 antibody
      • Heat shock protein 90 kDa antibody
      • Heat shock protein 90kDa alpha (cytosolic) class B member 1 antibody
      • Heat shock protein 90kDa alpha family class B member 1 antibody
      • Heat shock protein beta antibody
      • Heat shock protein HSP 90 beta antibody
      • Heat shock protein HSP 90-beta antibody
      • HS90B_HUMAN antibody
      • HSP 84 antibody
      • HSP 90 antibody
      • HSP 90 b antibody
      • HSP 90b antibody
      • HSP84 antibody
      • HSP90 BETA antibody
      • hsp90ab1 antibody
      • HSP90B antibody
      • HSPC2 antibody
      • HSPCB antibody
      see all

    Images

    • Western blot - Anti-Hsp90 beta antibody [H90-10] (ab53497)
      Western blot - Anti-Hsp90 beta antibody [H90-10] (ab53497)
      All lanes : Anti-Hsp90 beta antibody [H90-10] (ab53497) at 1/5000 dilution

      Lane 1 : Wild-type HEK-293T cell lysate
      Lane 2 : HSP90AB1 knockout HEK-293T cell lysate
      Lane 3 : Saos-2 cell lysate
      Lane 4 : HL-60 cell lysate

      Lysates/proteins at 20 µg per lane.

      Performed under reducing conditions.

      Predicted band size: 83 kDa
      Observed band size: 85 kDa why is the actual band size different from the predicted?



      Lanes 1 - 4: Merged signal (red and green). Green - ab53497 observed at 85 kDa. Red - loading control ab181602 (Rabbit Anti-GAPDH antibody [EPR16891]) observed at 37kDa.

      ab53497 was shown to react with Hsp90 beta in wild-type HEK-293T cells in western blot with loss of signal observed in HSP90AB1 knockout cell line ab266117 (HSP90AB1 knockout cell lysate ab257190). Wild-type and HSP90AB1 knockout HEK-293T cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% milk in TBS-T (0.1% Tween®) before incubation with ab53497 and ab181602 (Rabbit Anti-GAPDH antibody [EPR16891]) overnight at 4°C at a 1 in 5000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Mouse IgG H&L (IRDye® 800CW) preabsorbed (ab216772) and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed (ab216777) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Hsp90 beta antibody [H90-10] (ab53497)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Hsp90 beta antibody [H90-10] (ab53497)
      Ab53497 staining Human normal placenta. Staining is localized to cytoplasmic compartment.
      Left panel: with primary antibody at 2 ug/ml. Right panel: isotype control.
      Sections were stained using an automated system DAKO Autostainer Plus , at room temperature. Sections were rehydrated and antigen retrieved with the Dako 3-in-1 AR buffers EDTA pH 9.0 in a DAKO PT Link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 minutes. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 minutes and detected with Dako Envision Flex amplification kit for 30 minutes. Colorimetric detection was completed with Diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that for manual staining we recommend to optimize the primary antibody concentration and incubation time (overnight incubation), and amplification may be required.
    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Hsp90 beta antibody [H90-10] (ab53497)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Hsp90 beta antibody [H90-10] (ab53497)

      Immunohistochemistry analysis of formalin-fixed paraffin-embedded inflamed mouse colon using ab53497 at 1/10000. Primary antibody was incubated for 12 hours at 4°C. Secondary Antibody was a Biotin Goat Anti-Mouse at 1/2000 dilution incubated for 1 hour at room temperature. Counterstain was Mayer Hematoxylin (purple/blue) nuclear stain at 200 µl for 2 minutes at room temperature. Magnification: 40x.

    • Immunocytochemistry/ Immunofluorescence - Anti-Hsp90 beta antibody [H90-10] (ab53497)
      Immunocytochemistry/ Immunofluorescence - Anti-Hsp90 beta antibody [H90-10] (ab53497)
      ICC/IF image of ab53497 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 (ab53497, 1µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-mouse IgG (H+L) used at a 1/1000 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.

    • Western blot - Anti-Hsp90 beta antibody [H90-10] (ab53497)
      Western blot - Anti-Hsp90 beta antibody [H90-10] (ab53497)
      Anti-Hsp90 beta antibody [H90-10] (ab53497) at 1 µg/ml + Recombinant human Hsp90 beta protein (Active) (ab80033) at 0.1 µg

      Secondary
      Goat Anti-Mouse IgG H&L (HRP) preadsorbed (ab97040) at 1/5000 dilution

      Developed using the ECL technique.

      Performed under reducing conditions.

      Predicted band size: 83 kDa


      Exposure time: 8 minutes
    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Hsp90 beta antibody [H90-10] (ab53497)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Hsp90 beta antibody [H90-10] (ab53497)

      Paraffin-embedded mouse backskin (epidermis) tissue fixed with Bouin's fixative, stained for Hsp90 beta using ab53497 at 1/100 dilution in immunohistochemical analysis. Primary antibody was incubated for 1 hour at room temperature. Secondary antibody was a FITC-conjugated goat anti-mouse (green) at 1/50 dilution ncubated for 1 hour at room temperature.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Hsp90 beta antibody [H90-10] (ab53497)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Hsp90 beta antibody [H90-10] (ab53497)

      Formalin-fixed, paraffin-embedded human colon carcinoma tissue stained for Hsp90 beta using ab53497 at 1/10000 in immunohistochemical analysis. Primary antibody was incubated for 12 hours at 4°C. Secondary antibody was an Alexa Fluor® 555 goat anti-mouse (red) at 1/5000 dilution incubated for 1 hour at room temperature.

      40x magnification.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Hsp90 beta antibody [H90-10] (ab53497)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Hsp90 beta antibody [H90-10] (ab53497)

      ab53497 at 100,000 dilution staining Hsp90 in human colon cancer tissue section by immunohistochemistry (Formalin/ PFA fixed paraffin-embedded tissue sections). A antibody amplifier™ system was used for staining. A HRP-conjugated secondary antibody was used at 1/10 dilution

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Hsp90 beta antibody [H90-10] (ab53497)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Hsp90 beta antibody [H90-10] (ab53497)

      ab53497 at 100,000 dilution staining Hsp90 beta in mouse colon tissue section by immunohistochemistry (Formalin/ PFA fixed paraffin-embedded tissue sections). A antibody amplifier™ system was used for staining. An Alexa Fluor®  568 conjugated secondary antibody was used at 1/10 dilution.

    • Western blot - Anti-Hsp90 beta antibody [H90-10] (ab53497)
      Western blot - Anti-Hsp90 beta antibody [H90-10] (ab53497)
      All lanes : Anti-Hsp90 beta antibody [H90-10] (ab53497)

      Lane 1 : Hsp90 beta protein
      Lane 2 : Hsp90 alpha protein

      Lysates/proteins at 2 µg per lane.

      Predicted band size: 83 kDa

    • Flow Cytometry - Anti-Hsp90 beta antibody [H90-10] (ab53497)
      Flow Cytometry - Anti-Hsp90 beta antibody [H90-10] (ab53497)
      Overlay histogram showing HeLa cells stained with ab53497 (red line). The cells were fixed with 80% methanol (5 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 (ab53497, 0.5µg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG2a [ICIGG2A] (ab91361, 1µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed.

    References (20)

    Publishing research using ab53497? Please let us know so that we can cite the reference in this datasheet.

    ab53497 has been referenced in 20 publications.

    • Khaledian B  et al. Inhibition of heat shock protein 90 destabilizes receptor tyrosine kinase ROR1 in lung adenocarcinoma. Cancer Sci 112:1225-1234 (2021). PubMed: 33370472
    • Liu Y  et al. Proteomic Analysis Revealed the Characteristics of Key Proteins Involved in the Regulation of Inflammatory Response, Leukocyte Transendothelial Migration, Phagocytosis, and Immune Process during Early Lung Blast Injury. Oxid Med Cell Longev 2021:8899274 (2021). PubMed: 34007409
    • Kovanicová Z  et al. Cold Exposure Distinctively Modulates Parathyroid and Thyroid Hormones in Cold-Acclimatized and Non-Acclimatized Humans. Endocrinology 161:N/A (2020). PubMed: 32242612
    • Snigireva AV  et al. A 2,5-Dihydroxybenzoic Acid-Gelatin Conjugate Inhibits the Basal and Hsp90-Stimulated Migration and Invasion of Tumor Cells. J Funct Biomater 11:N/A (2020). PubMed: 32503118
    • Si F  et al. Porcine Epidemic Diarrhea Virus ORF3 Protein Is Transported through the Exocytic Pathway. J Virol 94:N/A (2020). PubMed: 32554695
    View all Publications for this product

    Customer reviews and Q&As

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    1-3 of 3 Abreviews or Q&A

    Western blot abreview for Anti-Hsp90 beta antibody [H90-10]

     Excellent
    Abreviews
    Application
    Western blot
    Sample
    Fish Tissue lysate - whole (Heart, brain, liver, white muscle, red muscle)
    Loading amount
    60 µg
    Specification
    Heart, brain, liver, white muscle, red muscle
    Gel Running Conditions
    Non-reduced Non-Denaturing (Native) (7.5)
    Blocking step
    Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
    Read More

    Dr. Katja Anttila

    Verified customer

    Submitted Mar 15 2011

    Question

    Thank your for your kindness,and I wish to have my antibodies refunded. Could you please tell 51AB company to contact me before refunding me?               

    Read More
    Answer

    Thank you for your reply. I will forward your request to 51AB, who will be in contact with you to process your refund request. I hope this information is helpful.  Please do not hesitate to contact us if you have any additional questions.

    Read More

    Question

    To whom it may concern, I bought two primary antibodies ab79849 and ab53497,which have been used in CoIP,but the results are confusing. I intend to verify the combination between HSP90α and HBP21,and that between HSP90β and HBP21using protein A+G agarose from Santa Cruz,and the results are as follows, If the  HBP21 antibody is used to pull down the protein complex,and tested respectively by HSP90 and HBP21 antibodies the band for HSP90 will show,which indicates the combination of HSP90 with HBP21. But if either of  those two HSP90 antibodies is used to pull down the protein complex,and tested respectively by HSP90 and HBP21 antibodies, there will be no band from HSP90,which is confusing,because both of your these two antibodies are applicable for IP as your website shows,which is inconsistent with my results.  

    Read More
    Answer

    Than you for your inquiry and I am sorry to hear that you are experiencing some trouble with ab53497 and ab79849. Are you running any isotype controls with these experiments? I just want to make sure that the pull down with the HBP21 antibody is genuine. Have you tried to do a regular IP and not a co-IP with ab53497 and ab79849? These are the IP references we know of with the HSP beta antibody, ab53497. Barent R. L. (1998) Mol. Endocrinol. 12: 342-354 11. Lo. M.A. (1998) EMBO J. 17: 6879-6887. What kind of lysis buffer are you using? A key factor in maintaining complex formation throughout the steps required for co-IP is the lysis and wash buffers. Many protein interactions will remain intact after lysis using standard non-denaturing lysis buffers. Buffers with low ionic strength (i.e., <120mM NaCl) that contain non-ionic detergents (NP-40 and Triton X-100) are less likely to disrupt protein-protein interactions; however, testing may be required to determine the best buffer formulation for a specific protein complex of interest. Additionally, lysing cells by sonication or vortexing the lysate or bead-bound immune complexes during the wash steps should be avoided to prevent the disruption of the protein-protein interaction(s) of the target complex. And while centrifugation is a standard method to separate the precipitated complexes from the remaining lysate and during wash steps, the samples should be handled gently to prevent the loss of bound complex proteins. An advanced technique to strengthen protein-protein interactions is by crosslinking the binding partners. Using this approach, all proteins within the active distance of the specific reagent in a cell lysate are covalently crosslinked, and the target protein can then be immunoprecipitated along with the other proteins in the complex without the risk of losing binding partners. Co-IPs can be tricky due to the nature of the interaction of the proteins, nonspecific binding to IP components and antibody contamination that may mask detection. We haven't tested ab53497 and ab79849 by co-IP so it's difficult to know if this may be the issue. Could you try the protocol recommendations or run a regular IP with the antibody? That could show that it is an issue with the protein binding more than the antibodies.

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