Anti-DHCR7 antibody (ab103296)

Submit a review Q&A (9)References (5)

Overview

  • Product name

    Anti-DHCR7 antibody
    See all DHCR7 primary antibodies

  • Description

    Rabbit polyclonal to DHCR7

  • Host species

    Rabbit

  • Tested applications

    Suitable for: ICC/IF, WBmore details

  • Species reactivity

    Reacts with: Mouse, Rat, Human

  • Immunogen

    Fragment protein corresponding to amino acids 1-259 of Human DHCR7 (NP_001351.2).

  • Positive control

    • Rat brain tissue lysate, Mouse testis tissue lysate, HepG2 cell lysate IF/ICC: HeLa cell line.

Properties

Applications

Our Abpromise guarantee covers the use of ab103296 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 5 µg/ml.
WB 1/500 - 1/1000. Predicted molecular weight: 54 kDa.

Target

  • Relevance

    The DHCR7 gene encodes delta-7-sterol reductase (EC 1.3.1.21), the penultimate enzyme of mammalian sterol biosynthesis that converts 7-dehydrocholesterol (7-DHC) to cholesterol. This enzyme removes the C(7-8) double bond introduced by the sterol delta8-delta7 isomerases. In addition, its role in drug-induced malformations is known: inhibitors of the last step of cholesterol biosynthesis such as AY9944 and BM15766 severely impair brain development (Moebius et al., 1998 [PubMed 9465114]).[OMIM]

  • Cellular localization

    Endoplasmic reticulum membrane; Multi-pass membrane protein

  • Database links

    • Alternative names

      • 7 dehydrocholesterol reductase antibody
      • 7 DHC reductase antibody
      • Delta 7 dehydrocholesterol reductase antibody
      • Putative sterol reductase SR 2 antibody
      • SLOS antibody
      • Sterol delta 7 reductase antibody
      • Sterol Delta antibody
      see all

    Images

    • Immunocytochemistry/ Immunofluorescence - Anti-DHCR7 antibody (ab103296)
      Immunocytochemistry/ Immunofluorescence - Anti-DHCR7 antibody (ab103296)

      ICC/IF image of ab103296 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 (ab103296, 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.

    • Western blot - Anti-DHCR7 antibody (ab103296)
      Western blot - Anti-DHCR7 antibody (ab103296)
      Anti-DHCR7 antibody (ab103296) at 1/500 dilution + Rat brain tissue lysate at 50 µg

      Predicted band size: 54 kDa

    • Western blot - Anti-DHCR7 antibody (ab103296)
      Western blot - Anti-DHCR7 antibody (ab103296)
      Anti-DHCR7 antibody (ab103296) at 1/500 dilution + Mouse testis tissue lysate at 50 µg

      Predicted band size: 54 kDa

    • Western blot - Anti-DHCR7 antibody (ab103296)
      Western blot - Anti-DHCR7 antibody (ab103296)
      Anti-DHCR7 antibody (ab103296) at 1/500 dilution + HepG2 cell lysate at 50 µg

      Predicted band size: 54 kDa

    References

    This product has been referenced in:

    • Zhang P  et al. Intrinsic BET inhibitor resistance in SPOP-mutated prostate cancer is mediated by BET protein stabilization and AKT-mTORC1 activation. Nat Med 23:1055-1062 (2017). Read more (PubMed: 28805822) »
    • Pirhaji L  et al. Revealing disease-associated pathways by network integration of untargeted metabolomics. Nat Methods 13:770-6 (2016). Read more (PubMed: 27479327) »
    See all 5 Publications for this product

    Publishing research using ab103296? 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 review Submit a question

    1-9 of 9 Abreviews or Q&A

    Question

    Thank you for the prompt reply. I talked with my supervisors and they agree with wanting to get a refund. I really appreciate your help through this process.

    Read More
    Answer

    Thank you for your reply.

    Your credit note ID is*/.

    I am sorry that this antibody did not perform as stated on the datasheet and that we were not able to get it working for you. 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

    Question

    So I have done the western blot with the new/fresh antibody you sent me for DHCR7. I have some additional questions.

    1) Have you or anyone with your company tested your antibody with human DHCR7 (cells, tissue....)? Your technical sheet indicates that the Ab. work for human DHCR7. SO, you must have tested those with human tissue or cells. Can you provide a picture of a blot obtained with human cells or tissue?

    2) In the brochure for the antibody, the blot shows two bands slightly above the 50kDa standard. Do you know exactly what the two proteins are shown in the blot on the abcam website? The picture of my blot is posted below. There are many, many bands, some of which are stronger than the 55kDa bands. In brief, the antibodies still don't seem very specific. I would understand perhaps seeing bands below 50kDa, which could be due to DHCR7 protein fragmentation through the isolation process. However, there are many bands above the 50kDa marker as well. The darkest band from the positive control you sent is around 100kDa.

    3) the positive control is far from being as clean as your picture suggests. Can you give us details on exactly how your electrophoresis (and Western blot) was run to produce the 2-band only blot? Inparticular, what are the conditions for "neutralizing" non-specific binding?

    4) your technical sheet indicates that the positive control was prepared from testis according to a proprietary procedure. Any chance you could disclose some elements of your proprietary recipe so we could try it on our cells?

    I would like to add that our team has decades or experience with SDS-PAGE and Western blotting techniques. I hope that in the end, our efforts with this project will not lead us to conclude that your antibodies have not been fully tested before being marketed.

    Read More
    Answer

    Thank you for your reply.

    I am sorry that you are still experiencing problems with ab103296 in western blotting.

    To answer your questions:

    1 - There is already an image showing the antibody react with HepG2 cells (human liver hepatocellular carcinoma cell line) on the webpages for this antibody. It shows a signal band at 50kDa:

    https://www.abcam.com/DHCR7-antibody-ab103296.html



    2 - The double banding is only seen in the mouse testis lysate, not in the rat or human lysates. Therefore we believe that this extra band is mouse specific and may represent a uncharacterized isoform or splice variant. For the WB image you submitted, I am unsure as to what the 100kDa band is, but we could work on blocking conditions and also lowering the amount of protein loaded into the well, along with lowering the primary antibody concentration, to help clean up this signal.

    3 - For the WB protocol, not special electrophoresis conditions were used. Standard protocols (see attached protocols) were used.

    4 - Unfortunately, as the technique is proprietary, I am unable to give out any information on this. However, safe to say standard isolation and extraction techniques should also be as successful.

    I would be happy to continue troubleshooting with you, but as you are having so much difficulty with this antibody, I would like to offer you a full refund.

    Please let me know how you would like to continue.

    Read More

    Question

    After talking with my PI, we decided that we would like to take you up on your offer of trying a new vial of the DHCR7 antibody. Thank you for getting back to me so quickly and if you like, I would be more than happy to follow up with you on how the new antibody works out (hopefully with wonderful results).

    Read More
    Answer

    You should have the new vial of ab103296 to try out tomorrow morning. Fingers crossed that it works for you. If not, please let me know (or if does work, I would like to know aswell).

    Hope you are having a great day.

    Read More

    Question

    Thank you for your input for blocking my membranes and helping me try to figure out using the antibody for DHCR7. I thought I would email you a follow up with how the positive control mouse testis lysate worked with the antibody…

    There still seems to be a lot of non-specific binding of the antibody as seen on the lanes with the positive control. Also, I am not sure of what to make out of the many more bands I see on the western done 7/2/12 other than possibly that the 7/2 wb picture is at a 5 minute exposure and the 6/21 wb picture is at a 3 min exposure. Are you sure that the western blot pictures on your website for DHCR7 antibody are accurate?

    Read More
    Answer

    Thank you for your reply and for sending those western blot images.

    I am very sorry that you are having issues with this antibody and I cannot think of anything else that would help resolve this issue. It could be that something happened to that particular vial that you received.

    I could either send a new vial of this antibody or I could process a refund. Please let me know how you would like to continue.

    Once again, I am sorry about the issues you have been having and I look forward to your reply and resolving this to your satisfaction.

    Enjoy the 4th.

    Read More

    Question


    Trying out your positive control would be wonderful! The purchase order for ab103296, is PO:1*****with the order number:*****. Also, I was wondering what the conditions were for your gel? where they reducing or non-reducing?

    Read More
    Answer

    Thank you for your reply.

    I am going to send the tissue lysate for you to use as a positive control in your western blots. The lysate is currently out of stock, but we are due to get it in on Friday (15th), so that means we should hopefully get it to you but Monday/Tuesday next. I am sorry about this delay and hopefully it will not delay your experiments for too long. Also the gels were run under reduced, denaturing conditions.

    If there is anything else I can help you with, please let me know.

    Read More

    Question


    I have attached the procedures I used and pictures of the gels and westerns. I appreciate your quick reply back to help me, I am under a time limit with this research.

    Read More
    Answer

    Thank you for your reply.

    Based on the information that you have provided, I think that the method of tissue preparation is fine, I don’t think that could be the issue. One piece of protocol that is different between what you do and the protocol that was used to test this antibody, is that in the protocol we have 5% non-fat dried milk. Switching to blocking agent could make a big difference.

    Also, as I said, I do not think you method of tissue preparation is the issue, to rule that out, I would be more than happy to send you one of our lysates to use as a positive control. I was thinking of:

    https://www.abcam.com/Testis-Mouse-Whole-Cell-Lysate-normal-tissue-ab29289.html



    If you would like me to send this sample as a positive control, please could you send me the Abcam order number for ab103296 and I will be happy to get this sent to you as soon as possible.

    I look forward to your reply.

    Read More

    Question

    There is a lot of none-specific binding of this primary antibody to proteins it should not be binding to (as in 15-20 bands at various molecular weights). In the western blot on the product handout there is only two bands. How did you isolate DHCR7 for the western blot you show, how did you run your western blot, and are there any references of other labs using your antibody?

    Read More
    Answer

    Thank you for contacting Abcam.

    I am sorry about the issues you have been having with ab103296. The antibody is covered under our Abpromise for six months and is guaranteed to work in western blot on Mouse, Rat and Human samples. If we cannot resolve the issue you are having with the antibody then I would be happy to either send a replacement antibody or to process a refund.

    To be able to help you further with this enquiry, would you please be able to provide me, with a copy of the protocol you used to produce your cell lysate (including the type and species of the sample) but also the western blot protocol that you used?

    I look forward to your reply and helping resolve this issue.

    Read More

    Question

    My lab recently purchased this polyclonal antibody specific to the protein DHCR7 from your company. I have been having trouble getting the antibody to attach to the protein for detection and because DHCR7 is not commercially available there is no possible way for me to confirm if the antibody is attaching or not. On your website, you have an image of the western blot proving that the antibody successfully attached to the protein. I am wondering where you got the DHCR7 protein for this western blot?

    Read More
    Answer

    Thanky you for your enquiry. I'm sorry to hear the vial of ab103296 you received is giving you trouble. The western blots shown on the online datasheet for ab103296 illustrate detection of DHCR7 from Rat brain tissue lysate, mouse testis tissue lysate and HepG2 cell lysate. We do have these extracts available for sale as positive controls for your western blot. Here are some examples: HepG2 cell lysate: https://www.abcam.com/HepG2-Human-hepatocellular-liver-carcinoma-cell-line-Whole-Cell-Lysate-ab7900.html Rat brain tissue lysate https://www.abcam.com/Brain-Rat-Membrane-Lysate-normal-tissue-ab29473.html mouse testis tissue lysate https://www.abcam.com/Testis-Mouse-Whole-Cell-Lysate-normal-tissue-ab29289.html I hope this is helpful. Please contact me again if you have any further questions.

    Read More

    Question

    Hello, My name is M and I have a quick question about the protocol you use to prepare the antibodies for DHCR7. In the information online it says that the peptide you use is 1-259 amino acids long. What do you use to solublize the original peptide? Thank you!

    Read More
    Answer

    Thank you for your enquiry. I can confirm the immunogen for ab103296 Anti-DHCR7 antibody is DHCR7 full-length human protein (NP_001351.1, amino acids 1 ~ 259), and its expression system is mammalian cell. I can confirm this item is sourced externally, and regrettably they are not able to provide us with information regarding the solubilization of the immunogen. I am sorry we do not have all the details you require on this occasion. If you have any further questions, please do not hesitaet to contact us.

    Read More

    Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
    For licensing inquiries, please contact partnerships@abcam.com