Anti-Firefly Luciferase antibody (ab21176)

Reviews (17)Q&A (18)References (45)

Overview

  • Product name

    Anti-Firefly Luciferase antibody
    See all Firefly Luciferase primary antibodies

  • Description

    Rabbit polyclonal to Firefly Luciferase

  • Host species

    Rabbit

  • Tested applications

    Suitable for: ICC/IF, IHC-Fr, WBmore details

  • Species reactivity

    Reacts with: Firefly

  • Immunogen

    Full length native protein (purified) (Firefly (Photinus pyralis)).

  • Positive control

    • ICC/IF: Firefly Luciferase transfected HEK-293 cells. Adult rat stromal cells. HEK-293 cells. Mouse mammary carcinoma cells. WB: Lysates from HEK-293T cells overexpressing luciferase. IHC-Fr: Mouse cerebellum tissue.

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: 0.097% Sodium azide
    Constituent: PBS
  • Concentration information loading...

  • Purity

    IgG fraction

  • Clonality

    Polyclonal

  • Isotype

    IgG

  • Research areas

Applications

Our Abpromise guarantee covers the use of ab21176 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 a concentration of 10 µg/ml.
IHC-Fr 1/1000. PubMed: 18219389
WB 1/1000 - 1/2000.

Target

  • Relevance

    Luciferase from the firefly has become one of the more widely used reporter proteins for the study of gene expression. Luciferase catalyzes a bioluminescent reaction which requires the substrate luciferin as well as Mg2+ and ATP. Mixing these reagents with the cell extract containing luciferase, results in a flash of light that decays rapidly. This light can be detected by a luminometer. The total light emission is proportional to the luciferase activity of the sample.

  • Cellular localization

    Peroxisome

  • Database links

    • Alternative names

      • ec 1.13.12.7 antibody
      • Firefly antibody
      • Luciferase antibody
      • Luciferin 4 monooxygenase antibody

    Images

    • Immunocytochemistry/ Immunofluorescence - Anti-Firefly Luciferase antibody (ab21176)
      Immunocytochemistry/ Immunofluorescence - Anti-Firefly Luciferase antibody (ab21176)

      ab21176 at 10 µg/mL staining Luciferase in transfected HEK-293 (Human epithelial cell line from embryonic kidney) cells by ICC/IF.

      The cells were fixed with methanol and acetone. An FITC conjugated anti-Rabbit IgG was used as the secondary antibody. 

      Left panel: Un-transfected cells.

      Right panel: Transfected cells.

    • Immunocytochemistry/ Immunofluorescence - Anti-Firefly Luciferase antibody (ab21176)
      Immunocytochemistry/ Immunofluorescence - Anti-Firefly Luciferase antibody (ab21176)This image is courtesy of an Abreview submitted by Dr Mal Niladri

      ab21176 at 1/200 dilution staining engineered adult rat stromal stem cells by ICC/IF.

      The cells were fixed in 2% paraformaldehyde and 0.1% Triton X-100 was used for cell permeabilization (15 minutes incubation time). The cells were incubated with the antibody overnight at 4°C. The image shows Luciferase (green-upper right panel), counterstained cell nuclei (DAPI-blue-upper left panel), overlay (lower left panel) and a phase contrast image (lower right panel).

      The image was taken with a confocal laser scanning microscope equipped with an additional laser differential Interference Contrast (DIC) mode.

      See Abreview

    • Immunohistochemistry (Frozen sections) - Anti-Firefly Luciferase antibody (ab21176)
      Immunohistochemistry (Frozen sections) - Anti-Firefly Luciferase antibody (ab21176)This image is courtesy of an Abreview submitted by Radhi Praba Velayutham

      ab21176 staining Firefly Luciferase in mouse cerebellum tissue sections by Immunohistochemistry (IHC-Fr - frozen sections).

      Tissue was fixed with acetone, permeabilized with PBS + 0.1% Triton X-100 (PBST) for 10 minutes and blocked with 10% serum for 1 hour at 22°C. Samples were incubated with primary antibody (1/1500 in 10% goat serum in PBST) for 1 hour at 22°C. An Alexa Fluor® 555-conjugated Goat anti-rabbit IgG polyclonal (1/1000) was used as the secondary antibody.

      See Abreview

    • Western blot - Anti-Firefly Luciferase antibody (ab21176)
      Western blot - Anti-Firefly Luciferase antibody (ab21176)
      All lanes : Anti-Firefly Luciferase antibody (ab21176) at 1/1000 dilution

      Lane 1 : Lysates from HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) cells overexpressing luciferase
      Lane 2 : Lysates from HEK-293T cells overexpressing luciferase with Luciferase Immunizing Peptide

      Secondary
      All lanes : Goat Anti-Rabbit IgG-Alkaline Phosphatase and a colorimetric substrate
    • Immunocytochemistry/ Immunofluorescence - Anti-Firefly Luciferase antibody (ab21176)
      Immunocytochemistry/ Immunofluorescence - Anti-Firefly Luciferase antibody (ab21176)

      Immunocytochemical analysis analysis of HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) cells overexpressing luciferase labeling Luciferase with ab21176 at a concentration of 10 μg/mL. The secondary antibody was a Goat anti-Rabbit IgG, FITC conjugate.

    • Immunocytochemistry/ Immunofluorescence - Anti-Firefly Luciferase antibody (ab21176)
      Immunocytochemistry/ Immunofluorescence - Anti-Firefly Luciferase antibody (ab21176)This image is courtesy of an anonymous Abreview

      ab21176 staining mouse mammary carcinoma cells by ICC/IF.

      Cells were PFA fixed and permeabilized in 0.1% Triton X-100 prior to blocking with a commercial blocking agent. The primary antibody was diluted 1/100 and incubated with the sample for 1 hour at 25°C. An Alexa-Fluor® 555 conjugated goat anti-rabbit antibody was used as the secondary.

      The image shows firefly luciferase (red) in mouse tumour cells and cell nuclei counterstained with Hoechst (blue).

      See Abreview

    References

    This product has been referenced in:

    • Fritzell K  et al. Sensitive ADAR editing reporter in cancer cells enables high-throughput screening of small molecule libraries. Nucleic Acids Res 47:e22 (2019). Read more (PubMed: 30590609) »
    • Kim S  et al. Optimizing live-animal bioluminescence imaging prediction of tumor burden in human prostate cancer xenograft models in SCID-NSG mice. Prostate 79:949-960 (2019). Read more (PubMed: 30958914) »
    See all 45 Publications for this product

    Publishing research using ab21176? Please let us know so that we can cite the reference in this datasheet.

    Customer reviews and Q&As

    Show All Reviews Q&A
    Submit a question

    1-10 of 18 Q&A

    Question

    Do you have any negative control, untransfected ICC/IF staining images? I'm interested to see any background signal.

    Read More
    Answer

    Thank you for your inquiry and your patience with this reply.

    After some further digging, the lab was able to provide a PPT of the ICC/IF results of untransfected vs transfected HEK293 cells stained using ab21176. The protocol is also included.

    I hope that this helps and please do not hesitate to contact us if you require any further information or assistance.

    Read More

    Question

    We ordered the antibody below for immunostaining of formalin-fixed, paraffin-embedded prostate samples from transgenic mice. Luciferase activity assay showed the prostate samples from transgenic mice express luciferase.    However, according to the protocol listed in Abcam, the prostate samples from both wild type (negative control) and transgenic mice have strong staining with no difference even at 1:1000 dilution of the antibody.  

    Read More
    Answer

    Thank you for contacting us. Your credit note ID is xxxx. I am sorry that this antibody did not perform as stated on the datasheet. I have asked our accounting department to issue a credit note for you, which can be redeemed against the invoice of a future order by passing it on to your purchasing department. To avoid confusion, please ensure your accounts department is aware of how the credit note is being used. If you have questions on how to use the credit note, please contact our accounting department. 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.

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    Question

    Dear Sir/Madam,   We ordered the antibody below for immunostaining of formalin-fixed, paraffin-embedded prostate samples from transgenic mice. Luciferase activity assay showed the prostate samples from transgenic mice express luciferase.    However, according to the protocol listed in Abcam, the prostate samples from both wild type (negative control) and transgenic mice have strong staining with no difference even at 1:1000 dilution of the antibody.   Can we replace another luciferase antibody from Abcam?

    Read More
    Answer

    Thank you for contacting us.





    I am sorry that this product has not performed as should be expected. This product is covered by our Abpromise guarantee. We are happy provide scientific support at any time. If you are using the product in applications listed on the datasheet and contact us within six months of purchase, we are also happy to replace or refund the product should we not be able to help you to resolve the issue.

    Would you be able to send me the details of your protocol so that I may get a better understanding of the problemms which you are facing. These details are also important should we decide that this product needs further internal testing.

    Could you also provide the Abcam order confirmation number or PO used?



    Thank you for your cooperation, I look forward to your response. 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

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    Question

    Do you have any negative control, untransfected ICC/IF staining images? I'm interested to see any background signal.

    Read More
    Answer

    Thank you for your inquiry.

    I heard back from the testing lab that they did not have any cell staining data for ab21176 with untransfected cells.

    I can point you to our references section of the online datasheet as you may be able to find one in a publication:

    https://www.abcam.com/Firefly-Luciferase-antibody-ab21176-references.html

    I hope this information helps. Please contact us with any other questions.

    Read More

    Question

    I had ordered for an
    anti-luciferase antibody, Ab21176, for the purpose of
    immunofluorescence staining on formalin fixed slides. Unfortunately,
    this has not worked for me too well. I would like to probe for my
    protein of interest using Western Blot instead. Can I use this same
    antibody? If yes do you have recommendations on the dilution?

    Read More
    Answer

    Thank you for contacting Abcam regarding ab21176.


    I am sorry that you were experiencing difficulties with this antibody in IHC. If you believe this is due to the antibody being defective, I am happy to provide additional technical assistance with troubleshooting or a credit, as this product is guaranteed for IF under our Abpromise. Please provide some additional protocol information.


    Regarding using the antibody in WB, we are not sure if this antibody will be suitable for western, but would recommend a starting dilution of 1 ug/ml.


    I hope this information is helpful. Please do not hesitate to contact me if you have any additional questions or concerns.

    Read More

    Question

    Name: Technical Service INTERLAB CO., LTD.
    Product code: 21176

    Inquiry: Dear technical support team: This customer raises an enquiry about ab21176 (Anti-Firefly Luciferase antibody), she wants to know is there any suitable positive control of this antibody (conduct WB)? Would you please help him to solve the problem? Thanks for your kindly help. Best regards Ada

    Read More
    Answer

    Thank you for contacting us.

    The customer could use lysates of HEK-293T cells overexpressing luciferase as positive control for ab21176.

    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

    Question

    Thank you for your prompt reply. Our order # is xxxxx.   In response to your email, I attached the protocol and the staining result. Please let us know if you need any further information.  

    Read More
    Answer

    Thank you for taking time to send me these details and for contacting us. I am sorry to hear this antibody is not providing satisfactory results.

    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, 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 am pleased to offer you a free of charge replacement, credit note, or refund in compensation.

    Thank you for your cooperation. I look forward to hearing from you with details of how you would like to proceed.

    Read More

    Question

    We see high background in nuclei following protocol suggested on the datasheet.

    Read More
    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.

    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.

    Read More

    Question

    then what secondary antibody can be used for ab64569?
    thanks

    Read More
    Answer

    Thank you for your enquiry.

    Here is an example of a secondary antibody you can use to detect ab64569:

    https://www.abcam.com/Goat-polyclonal-Secondary-Antibody-to-Mouse-IgG-Fc-DyLight-488-pre-adsorbed-ab98712.html

    ab98712 is conjugated to a fluor that can be visualized with the same filter set as is used for FITC. This antibody has been preadsorbed to rat serum (among others) and has shown minimum cross reactivity to rat immunoglobulins.

    We have a couple useful tools on our website for selecting secondary antibodies. One is a search and another is a browse feature and both can be found at:
    https://www.abcam.com/index.html?pageconfig=productmap&cl=918

    I hope this is helpful. Please contact me again if you have any further questions.

    Read More

    Question

    Hi
    I have decided to order the following primary antibodies:
    https://www.abcam.com/Endostatin-antibody-4i37-ab64569.html (to detect endostatin, expressed by the gene carriers in rat cornea)
    https://www.abcam.com/Firefly-Luciferase-antibody-Luci17-ab16466.html (to detect luciferase, expressed by the gene carriers in rat cornea)
    Both cornea samples were PFA fixed and parrafin blocks. The I am thinking to order secondary antibody http://https://www.abcam.com/Goat-polyclonal-Secondary-Antibody-to-Mouse-IgG1-heavy-chain-FITC-ab97239.html
    Could you please give me technical support about my selection? Especially how secondary antibody to match my primary antibody? Since the proteins we are going to detects are in rat, I just wonder if the one I selected is fine. Do you have any good suggestions.
    I am going to use Zeiss 710 confocal, filters are no problem for my research.

    Read More
    Answer

    Thank you for your email.

    ab97239 is specific for the heavy chain of mouse IgG1. The subtype for ab64569 is not known. If this antibody is not IgG1 then ab97239 will not bind. The subtype of ab16466 is IgG1, so ab97239 will bind this IgG heavy chain.

    I hope this is helpful. Please contact me again if you have any further questions.

    Read More

    1-10 of 18 Q&A

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