Overview

  • Product name

    Anti-Iba1 antibody [1022-5]
    See all Iba1 primary antibodies
  • Description

    Mouse monoclonal [1022-5] to Iba1
  • Host species

    Mouse
  • Specificity

    Some customers have reported successful staining using this antibody on frozen sections, however ab15690 is not routinely tested on frozen sections and we do not recommend this antibody for use with frozen tissue. In our hands this product worked well in WB under standard reducing and denaturing conditions (please see image below). ab15690 detected Iba1 migrating at approximately 17 kDa in positive control samples (Rat spleen, THP1, C6) and bound no protein in samples we believe have little or no Iba1 expression (HEK293, NIH3T3 and A431). We recommend blocking in milk. Blocking with BSA gives high background.
  • Tested applications

    Suitable for: ICC/IF, WB, IHC-Pmore details
    Unsuitable for: IHC-Fr
  • Species reactivity

    Reacts with: Rat, Human
  • Immunogen

    Recombinant full length protein (Human).

  • Positive control

    • Spleen, THP1, C6

Properties

Applications

Our Abpromise guarantee covers the use of ab15690 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 Use at an assay dependent concentration. PubMed: 24916922
WB Use a concentration of 1 µg/ml. Detects a band of approximately 17 kDa (predicted molecular weight: 17 kDa).

We recommend blocking in milk. Blocking with BSA gives high background.

IHC-P Use a concentration of 10 - 20 µg/ml. Perform heat mediated antigen retrieval via the microwave method before commencing with IHC staining protocol.
  • Application notes
    Is unsuitable for IHC-Fr.
  • Target

    • Function

      Actin-binding protein that enhances membrane ruffling and RAC activation. Enhances the actin-bundling activity of LCP1. Binds calcium. Plays a role in RAC signaling and in phagocytosis. May play a role in macrophage activation and function. Promotes the proliferation of vascular smooth muscle cells and of T-lymphocytes. Enhances lymphocyte migration. Plays a role in vascular inflammation.
    • Tissue specificity

      Detected in T-lymphocytes and peripheral blood mononuclear cells.
    • Sequence similarities

      Contains 2 EF-hand domains.
    • Post-translational
      modifications

      Phosphorylated on serine residues.
    • Cellular localization

      Cytoplasm > cytoskeleton. Cell projection > ruffle membrane. Associated with the actin cytoskeleton at membrane ruffles and at sites of phagocytosis.
    • Information by UniProt
    • Database links

    • Alternative names

      • AIF 1 antibody
      • AIF-1 antibody
      • Aif1 antibody
      • AIF1 protein antibody
      • AIF1_HUMAN antibody
      • Allograft inflammatory factor 1 antibody
      • Allograft inflammatory factor 1 splice variant G antibody
      • allograft inflammatory factor-1 splice variant Hara-1 antibody
      • balloon angioplasty responsive transcription antibody
      • BART 1 antibody
      • G1 antibody
      • G1 putative splice variant of allograft inflamatory factor 1 antibody
      • IBA 1 antibody
      • IBA1 antibody
      • interferon gamma responsive transcript antibody
      • Interferon responsive transcript 1 antibody
      • interferon responsive transcript factor 1 antibody
      • Ionized calcium binding adapter molecule 1 antibody
      • Ionized calcium-binding adapter molecule 1 antibody
      • ionized calcium-binding adapter molecule antibody
      • IRT 1 antibody
      • IRT1 antibody
      • Microglia response factor antibody
      • MRF1 antibody
      • Protein g1 antibody
      see all

    Images

    • IHC image of Iba1 staining in human normal spleen formalin fixed paraffin embedded tissue section*, performed on a Leica Bond™ system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab15690, 10µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

       

      For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

      *Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

    • All lanes : Anti-Iba1 antibody [1022-5] (ab15690) at 1 µg/ml

      Lane 1 : Human Iba1 full length recombinant protein at 0.1 µg
      Lane 2 : HEK293 whole cell lysate at 20 µg
      Lane 3 : A431 whole cell lysate at 20 µg
      Lane 4 : NIH3T3 whole cell lysate at 30 µg
      Lane 5 : Human spleen tissue lysate at 20 µg
      Lane 6 : Mousespleen tissue lysate at 30 µg
      Lanes 7 & 16 : Rat spleen tissue lysate at 30 µg
      Lane 8 : U937 whole cell lysate at 20 µg
      Lane 9 : MOLT4 whole cell lysate at 20 µg
      Lanes 10 & 17 : THP1 whole cell lysate at 30 µg
      Lane 11 : THP1 whole cell lysate, PMA treated at 30 µg
      Lane 12 : Raw 264.7 whole cell lysate at 30 µg
      Lane 13 : C6 whole cell lysate at 30 µg
      Lane 14 : NR8383 whole cell lysate at 30 µg
      Lane 15 : Mouse spleen tissue lysate at 30 µg

      Developed using the ECL technique.

      Performed under reducing conditions.

      Predicted band size: 17 kDa



      Lanes 1-14: Blocked in 3% milk for 1 hour (RT). Lanes 15-17: Blocked in 5% BSA for 1 hour (RT).

      Lane 1: exposure 1 minute. Lanes 2-17: exposure 8 minutes. Abcam recommends blocking in milk for cleaner blots with reduced background, in comparison to BSA. 

      This blot was produced using a 4-12% Bis-Tris gel under the MOPS buffer system. The gel was run at 200V for 60 minutes before being transferred onto a nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour before being incubated with ab15690 (anti-Iba1 antibody; 1 ug/mL) for 18 hours at 4°C. Antibody binding was detected using ab97040 (HRP-labelled goat anti-mouse IgG) at  1:50,000 dilution for 1 hour at room temperature and visualised using ECL development solution ab133406.

    • Ab15690 staining IBA1 in human induced microglia by ICC/IF (Immunocytochemistry/Immunofluorescence). Cells were fixed with 4% paraformaldehyde for 15 minutes. Samples were incubated with primary antibody (1:200) for 2 hours at room temperature. Hoechst nuclear stain (blue) was used at 1:5000 dilution. 

    • Immunohistochemical analysis of paraffin embedded human speen sections, labelling Iba1 with ab15690.

    • Immunohistochemical analysis of paraffin embedded human speen sections, labelling Iba1 with ab15690.

    References

    This product has been referenced in:

    • Amal H  et al. S-nitrosylation of E3 ubiquitin-protein ligase RNF213 alters non-canonical Wnt/Ca+2 signaling in the P301S mouse model of tauopathy. Transl Psychiatry 9:44 (2019). Read more (PubMed: 30696811) »
    • Hu JZ  et al. Silencing of lncRNA PKIA-AS1 Attenuates Spinal Nerve Ligation-Induced Neuropathic Pain Through Epigenetic Downregulation of CDK6 Expression. Front Cell Neurosci 13:50 (2019). Read more (PubMed: 30873006) »
    See all 77 Publications for this product

    Customer reviews and Q&As

    1-10 of 26 Q&A

    Answer

    This concentration is indeed quite high for an antibody so if you wish to dilute it in PBS then that is fine. I would say, that diluting it down to 1 mg/ml would be fine and is standard for most antibodies. Diluting it down to 100 ug/ml is quite a big dilution and I would not recommend this as it may affect the stability of this product especially when storing it at -20ºC or -80ºC.
    I notice that you are wanting to use this antibody for IHC-FrFl on rat brain tissue. As stated on the datasheet, although some customers have reported successful staining using this antibody on frozen sections, we do not routinely test this antibody on frozen sections, and we do not recommend it for use on these. For this reason, we have not suggested a dilution factor for this application. If you still want to proceed with this experiment I would suggest following the IHC-P guidelines so using 10-20 ug/ml of antibody per experiment.

    Read More

    Answer

    Thank you for your enquiry.

    I would like to reassure you that our antibodies have all been tested successfully and would be covered by our guarantee in the applications listed on the datasheets. Therefore, any antibody tested in the species and application you are using should be suitable for your applications.

    Also, although we produce many in-house antibodies, we obtain other antibodies from a wide range of sources. Therefore, comparison experiments will not often have been done.

    If you wish to be more certain, I can suggest choosing the antibody that has been best characterized. By this I mean the antibody that has been tested in the most applications and has the most data available (on the datasheet).

    In regard to your concerns about ab15690 Anti-Iba1 antibody [1022-5], we do offer another Anti-Iba1 which has been raised in goat and has a number of referenced publications, Abreviews and support data. I would recommend this product as an alternative to ab15690.

    https://www.abcam.com/iba1-antibody-ab5076.html


    I look forward to hearing from you. Should you have any further questions please do not hesitate to ask.

    Read More

    Question
    Answer

    Thank you for contacting us.

    Please find the information below describingour testing discount program:

    For UNTESTED species and/or applications, we have established a testing discount program. Here is a brief description of how it works:

    The testing discount program is for customers who like to use an antibody/protein/kit on an untested species/application. You would purchase the antibody/protein/kit at full price, test it and submit an Abreview with your data (positive or negative). On your next order you will receive a discount for ONE antibody/protein at the full price (100%) of the antibody/protein you have tested, or a full price discount for the amount paid for the kit. The terms and conditions applicable to this offer can be found here: https://www.abcam.com/collaborationdiscount.
    This programapplies to ELISA kits and other kits we offer.


    Also, I wanted to check homology betweenhuman and rat MMP1, but was not able to find the full length protein sequence for rat MMP1 in the database.
    However, if you like to try any of the antibodies that do not list "rat" as tested species (only as predicted), please let me know and I'd be happy to send you a testing discount code.

    I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

    Use our products? Submit an Abreview. Earn rewards!
    https://www.abcam.com/abreviews

    Read More

    Answer

    Thank you for your reply with the requested protocol information.


    I noticed there was no mention of any permeabilization or treatment with detergent. Have you tried adding Tween or Triton to TBS to perform a separate permeabilization or included this in your block or wash buffer? This would help the antibody to penetrate the tissue and increase the signal strength.


    If you find that this does not improve your results, I am happy to provide a credit. Please also provide an order number for ab15690 and ab102876.

    Read More

    Answer

    Thank you for contacting Abcam regarding ab15690.


    I have reviewed the history of this antibody and we have had some reports of high background in a recent lot. These complaints have been resolved by protocol modifications or replacements form a new lot.


    I am hoping you would send me a summary of your protocol as well as an image of the data you are obtaining with this antibody. I will then see if protocol suggestions may solve the problem or provide a replacement or credit.


    I look forward to your reply so that I may assist you further. Please do not hesitate to contact me if you have any additional questions.

    Read More

    Answer

    Thank you for taking time to complete our questionnaire and sending us further details.
    The details provided will enable us to investigate this case and will provide us with vital information for monitoring product quality. I appreciate the time you have spent in the laboratory and understand your concerns. It is regrettable the results have not been successful.
    Having reviewed the protocol details sent, I believe this product should have given satisfactory results. It appears that you may have received a faulty vial.
    I apologize for the inconvenience and have just raised a free of charge replacement with order number XXXXX. To check the status of the order please contact our Customer Service team and reference this number.
    You can visit our protocols online for tips and protocol information.
    I hope this new vial works as expected. Please do not hesitate to contact us if you need any more advice or information.

    Read More

    Question
    Answer

    Thank you for confirming these details and for your cooperation. The details provided enable us to closely monitor the quality of our products.

    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 with the order number ***.

    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 sending the details of your protocol. We usually recommend a permeabilization step with a solution of detergent when staining section thicker than 20 microns or so, but since the issue is non-specific staining, I am skeptical that this will help.

    If payment has already been made on the original order and you wish to receive a refund, please ask your purchasing department to contact our accounting department so that we may gather the correct information needed for the refund. To avoid confusion, please ensure your accounts department is aware of how the credit note is being used.

    Our accounting department can be contacted by email at us.credits@abcam.com or by telephone using the information at the Contact Us link in the top right corner of our website. Please refer to the credit note ID in any correspondence with our accounting department.

    The credit note ID is for your reference only and does not automatically guarantee the credit.

    I hope this experience will not prevent you from purchasing other products from us in the future. Our Scientific Support team is always at your service, should you require further expert advice.

    Read More

    Answer

    Thank you for brining this to our attention. Could you please tell me the purchase order number and approximate date of the order, or the Abcam order reference number, if you have it?

    We may be able to make some suggestions that improve your results. Can you please tell me how you prepared the brain tissue, in particular, how you fixed it and embedded it, and how thick the sections were? Were there any steps to the staining procedure aside from applying the antibody? Did you include a no-primary-antibody negative control to check for non-specific staining contributed by the secondary antibody?

    I look forward to your reply. If we cannot offer suggestions that improve your results, a refund will be provided.

    Read More

    Answer

    Thank you for taking the time to contact us. I am sorry to hearthe customer hashad difficulty obtaining satisfactory results from these threeantibodies.

    I would like to reassure you that in the event that a product is not functioning in the tested applications and species cited on the product data sheet (and the problem has been reported within 6 months of purchase), we will be pleased to provide a credit note, free of charge replacement or refund.

    I would like to investigate this particular case further for you, and also obtain more information for our quality monitoring records. In order to proceed with this, I have enclosed a technical questionnaire below. I would appreciate if you could complete this. It will help you put the information we require together very easily.

    I would appreciate if you could also provide images which would help us to assess the results.

    Please note Ihave sent the questionnaire for IHC and ICC. If the customer has used another application, please let me know and I can send the correct form so we can obtain the relevant information required.

    Thank you for your time and cooperation. We look forward to receiving the completed questionnaire.



    Order Details
    Antibody code:

    Lot number:

    Purchase order number
    or preferably Abcam order number:


    General Information
    Antibody storage conditions (temperature/reconstitution etc)


    Description of the problem (high background, low signal, non-specific satining etc.)

    Sample (Species/Tissue/Cell Type/Cell Line etc.)


    Fixation of sample (Ethanol/Methanol/Acetone/Paraformaldehyde/Other/Duration etc.)


    Antigen retrieval (Enzymatic method, Heat mediated technique etc.)


    Permeabilization step


    Blocking conditions (Buffer/time period, Blocking agent etc.)


    Primary Antibody (Manufacturer/Species/Diluent/Dilution/Incubation time, Wash step)


    Secondary Antibody (Manufacturer/Species/Diluent/Dilution/Incubation time, Wash step)


    Detection method


    Positive and negative controls used (please specify)


    Optimization attempts (problem solving)
    How many times have you tried the IHC?



    Have you run a "No Primary" control?
    Yes No

    Do you obtain the same results every time?
    Yes No


    What steps have you altered?


    Additional Notes


    We would appreciate if you are also able to provide and image which woudl help us to assess the results

    Read More

    1-10 of 26 Q&A

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