Overview

  • Product name
    Anti-BCRP/ABCG2 antibody [BXP-53]
    See all BCRP/ABCG2 primary antibodies
  • Description
    Rat monoclonal [BXP-53] to BCRP/ABCG2
  • Host species
    Rat
  • Tested applications
    Suitable for: ICC, WB, Flow Cyt, IHC-P, ICC/IFmore details
    Unsuitable for: EMSA
  • Species reactivity
    Reacts with: Mouse, Human
  • Immunogen

    Fusion protein containing the E. coli maltose binding protein and a fragment corresponding to amino acids 221-394 of Mouse BCRP/ABCG2

  • Epitope
    Reacts with an internal epitope of BCRP/ABCG2.

Properties

Applications

Our Abpromise guarantee covers the use of ab24115 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 1/20 - 1/50. Acetone fixed cytospin.
WB 1/20 - 1/50. Detects a band of approximately 75 kDa (predicted molecular weight: 72 kDa).
Flow Cyt Use 1-2µg for 106 cells.

(1/10 dilution on bone marrow and kidney tissue).

 

 

 

ab18450 - Rat monoclonal IgG2a, is suitable for use as an isotype control with this antibody.

 

IHC-P Use at an assay dependent concentration. PubMed: 20472681
ICC/IF Use at an assay dependent concentration.
  • Application notes
    Is unsuitable for EMSA.
  • Target

    • Function
      Xenobiotic transporter that may play an important role in the exclusion of xenobiotics from the brain. May be involved in brain-to-blood efflux. Appears to play a major role in the multidrug resistance phenotype of several cancer cell lines. When overexpressed, the transfected cells become resistant to mitoxantrone, daunorubicin and doxorubicin, display diminished intracellular accumulation of daunorubicin, and manifest an ATP-dependent increase in the efflux of rhodamine 123.
    • Tissue specificity
      Highly expressed in placenta. Low expression in small intestine, liver and colon.
    • Sequence similarities
      Belongs to the ABC transporter superfamily. ABCG family. Eye pigment precursor importer (TC 3.A.1.204) subfamily.
      Contains 1 ABC transmembrane type-2 domain.
      Contains 1 ABC transporter domain.
    • Post-translational
      modifications
      Glycosylation-deficient ABCG2 is normally expressed and functional.
    • Cellular localization
      Cell membrane.
    • Information by UniProt
    • Database links
    • Alternative names
      • ABC transporter antibody
      • ABC15 antibody
      • ABCG 2 antibody
      • ABCG2 antibody
      • ABCG2_HUMAN antibody
      • ABCP antibody
      • ATP binding cassette sub family G (WHITE) member 2 antibody
      • ATP binding cassette transporter G2 antibody
      • ATP-binding cassette sub-family G member 2 antibody
      • BCRP antibody
      • BCRP1 antibody
      • BMDP antibody
      • Breast cancer resistance protein antibody
      • CD338 antibody
      • CDw338 antibody
      • CDw338 antigen antibody
      • EST157481 antibody
      • GOUT1 antibody
      • MGC102821 antibody
      • Mitoxantrone resistance associated protein antibody
      • Mitoxantrone resistance-associated protein antibody
      • MRX antibody
      • Multi drug resistance efflux transport ATP binding cassette sub family G (WHITE) member 2 antibody
      • MXR antibody
      • MXR1 antibody
      • Placenta specific ATP binding cassette transporter antibody
      • Placenta specific MDR protein antibody
      • Placenta-specific ATP-binding cassette transporter antibody
      • UAQTL1 antibody
      see all

    Images

    • All lanes : Anti-BCRP/ABCG2 antibody [BXP-53] (ab24115) at 1 µg/ml

      Lane 1 : Caco 2 (Human colonic carcinoma cell line) Whole Cell Lysate
      Lane 2 : HepG2 (Human hepatocellular liver carcinoma cell line) Whole Cell Lysate

      Lysates/proteins at 10 µg per lane.

      Secondary
      All lanes : Peroxidase Conjugated AffiniPure Rabbit Anti-Rat IgG (H+L) at 1/10000 dilution

      Developed using the ECL technique.

      Performed under reducing conditions.

      Predicted band size: 72 kDa
      Observed band size: 75 kDa
      why is the actual band size different from the predicted?
      Additional bands at: 37 kDa. We are unsure as to the identity of these extra bands.


      Exposure time: 20 minutes


      BCRP/ABCG2 contains a potential glycosylation site (SwissProt) which may explain its migration at a higher molecular weight than predicted.
    • Overlay histogram showing HEK293 cells stained with ab24115 (red line). The cells were fixed with 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 (ab24115, 1µg/1x106 cells) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-rat IgG (Fc) (ab96971) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was rat IgG2a [aRTK2758] (ab18450, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a significantly decreased signal in HEK293 cells fixed with 4% paraformaldehyde (10 min)/permeabilized in 0.1% PBS-Tween used under the same conditions.

    • ab24115 staining BCRP/ABCG2 in murine mesenchymal stem cells by Immunocytochemistry/ Immunofluorescence. The cells were fixed in formaldehyde, permeabilised in 0.1% Triton X and then blocked using 1% serum for 1 hour at 25°C. Samples were then incubated with primary antibody at 1/25 for 15 hours at 25°C. The secondary antibody used was a donkey anti-rat IgG conjugated to an Alexa Fluor® used at a 1/100 dilution.

      See Abreview

    References

    This product has been referenced in:
    • Wang W  et al. SOX2OT variant 7 contributes to the synergistic interaction between EGCG and Doxorubicin to kill osteosarcoma via autophagy and stemness inhibition. J Exp Clin Cancer Res 37:37 (2018). WB ; Human . Read more (PubMed: 29475441) »
    • Zhang W  et al. Rab23 promotes the cisplatin resistance of ovarian cancer via the Shh-Gli-ABCG2 signaling pathway. Oncol Lett 15:5155-5160 (2018). Read more (PubMed: 29552151) »
    See all 26 Publications for this product

    Customer reviews and Q&As

    1-10 of 20 Abreviews or Q&A

    Abcam has not validated the combination of species/application used in this Abreview.
    Application
    Immunohistochemistry (PFA perfusion fixed frozen sections)
    Sample
    Rat Tissue sections (BRAIN)
    Antigen retrieval step
    Other
    Permeabilization
    Yes - TRITON 1%
    Specification
    BRAIN
    Blocking step
    Serum as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5 · Temperature: 20°C
    Fixative
    Paraformaldehyde

    Mrs. Francoise Geffroy

    Verified customer

    Submitted Jun 20 2018

    Answer

    Free of charge replacement being sent.

    Read More

    Answer

    Thank you for taking the time to contact us and provide the details. I am sorry to hear you have had difficulty obtaining satisfactory results from this antibody.

    The details you have kindly provided will enable us to investigate this case for you and this is also helpful in our records for monitoring of quality.

    I would like to reassure you that ab24115 is tested and covered by our 6 month guarantee for use in ICC-IF and human samples. In the event that a product is not functioning in the applications and species cited on the product data sheet, we will be pleased to provide a credit note or free of charge replacement.

    Reviewing this case, I would like to offer some suggestions to help optimize the results. I would also appreciate if you can confirm some further details:

    1. I am sorry to confirm this antibody is not tested in dog samples, it is tested and covered by the guarantee in human samples.

    2. Please confirm the order number and date of purchase

    3. How has the antibody been stored?

    4. I would appreciate if you are able to provide any images which would help me to assess the results.

    5. I can recommend to try overnight incubation 4oC. This can often provide more specific and efficient staining.

    6. Will the secondary antibody detect the rat IgG2a isotype? What is the supplier and product code? Has the current vial worked well with other rat IgG2a primary antibodies?

    7. Looking at the Abreviews on the datasheet, other customers have used acetone fixation successfully (which wil also permeabilize the cells). Others have fixed in paraformaldehyde then permeabilized in Triton. I can recommend to try this. It may be that the epitope is on the intracellular side of the membrane and so permeabilization is required.

    I hope this information is helpful, thank you for your cooperation. Should the suggestions not improve the results, please do not hesitate to contact me again with the further requested details.

    Read More

    Answer

    Thank you for your reply.

    I have to confirm that we did not test this antibody in IF with paraffin embedded sections.

    This is in theory possible. Although often high background occurs with this method.

    Please follow this link for a protocol:

    https://www.abcam.com/index.html?pageconfig=resource&rid=11459

    I can also recommend this page for in depth information about IHC:

    http://www.ihcworld.com/knowledge-center.htm

    I hope this information is helpful and wish you good luck with your research.

    Read More

    Answer

    Thank you for your inquiry.

    I am happy to confirm that both antibodies are tested and guarantee for IHC-P.

    1.) We used ab52816 successfully in a dilution of 1/100 in IHC-P with human squamous cervical carcinoma by Immunohistochemistry, Paraffin embedded tissue.

    2.) ab24115 was used at a dilution od 1/400 in the following publication:

    http://www.ncbi.nlm.nih.gov/pubmed/20472681?dopt=Abstract

    Please not that these dilution factors are only recommended starting points for your own optimization experiments.

    I wish you good luck with your experiments.

    Read More

    Answer

    I am sorry this product did not perform as stated on the datasheet and for the inconvenience this has caused. As requested, I have issued a free of charge replacement for one vial of ab24115.

    To check the status of the order please contact our Customer Service team and reference this number.

    Please note that this free of charge replacement vial is also covered by our Abpromise guarantee. Should you still be experiencing difficulties, or if you have any further questions, please do not hesitate to let us know.

    I wish you the best of luck with your research.

    Read More

    Answer

    Thank you for your reply. As we had previously discussed, ab24115 is not in an ideal formulation for use with our conjugation kits due to the fact that is not a purified antibody and it contains BSA as a stabilizer protein. I am sorry that these products were unable to produce good results in your experiment, and if you want I would be happy to either refund your kit purchase, or send you an alternative antibody to try as a free of charge replacement. Please let me know how you would like to proceed and I will be happy to assist you further.

    Read More

    Answer

    Thank you for contacting us.


    I have discussed the EMSA results with my colleagues, and to be honest we do not have any specificsuggestions:

    Because there are many unique requirements for each nucleic acid binding protein and the corresponding antibody, there is no universal set of reaction conditions. If no information on prior work is available, conditions should be determined empirically.In general, the supershift assay is a step which can be optimised in various ways. So could the incubation time and temperature be altered and may range from for example 1 hour to overnight at 4C or 15-45 min at room temperature. Or the antibody could be addedin different concentrations before or after the oligos. Also, factors that affect the strength and specificity of the protein:DNA interactions include the ionic strength and pH of the binding buffer, the presence of nonionic detergents, glycerol or carrier proteins (e.g., BSA),the concentration and type of competitor DNA present.

    If the customer has tried various conditions (which I am sure with her experience she has) and provided the ABCG2 protein construct is capable of binding to DNA, it might well be that the antibody is not suitable for EMSA. Antibodies being used to create a supershift may not work in a binding buffer that is optimal for a protein:DNA shift complex.Many nuclear proteins function well for shift complexes under reducing conditions. Therefore concentrations of DTT at 1 mM and higher are often added to the EMSA binding buffer. Yet, many antibodies become reduced and loose functionality in reducing conditions. Also, the epitope may not be in an appropriate location which preventsbinding of the antibody when the protein is bound to DNA.

    Therefore,the outcomeof theexperiments seems unfortunately negative, however, it is very good toknowunder which conditions the antibodydoes not work!

    Thank you very much for your time and contribution! I am sorry that I cannot be of more assistance on this occasion to help improve the results.

    Read More

    Answer

    Thank you for contacting Abcam. Unfortunately we do not have a blocking peptide available for this product. The sequence that was used to generate this antibody was ******** of Mouse BCRP/ABCG2 (SwissProt: Q7TMS5). You could have a peptide synthesized against this sequence and use that peptide for your blocking experiment. If there is anything else I can help you with, please let me know.

    Read More

    Answer

    DISCOUNT CODE: xxx Expiration date:dd mm 2012  I am very pleased to hear you would like to accept our offer and test ab24115 in EMSA. This code will give you 1 free primary antibody before the expiration date. To redeem this offer, please submit an Abreview for EMSA and include this code in the “Additional Comments” section so we know the Abreview is for this promotion. For more information on how to submit an Abreview, please visit the site: www.abcam.com/Abreviews. Remember, we publish both positive and negative Abreviews on our datasheets so please submit the results of your tests. The code will be active once the Abreview has been submitted and can be redeemed in one of the following ways: 1) Call to place your order and mention the code to our customer service department; 2) Include the code in your fax order; 3) Place your order on the web and enter the promotional code. Any feedback that you can provide will be greatly appreciated, whether positive or negative. If you have any further questions, please do not hesitate to contact us. We look forward to receiving your Abreview and wish you luck with your research. The terms and conditions applicable to this offer can be found here: www.abcam.com/collaborationdiscount.

    Read More

    1-10 of 20 Abreviews or Q&A

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

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