Anti-DMC1 抗体 [2H12/4] (ab11054)

Thank you for your enquiry.

I am sorry to hear the replacement antibody is still not working as expected. I would be happy to send you different Dmc1 antibody of your choice. So I can process your request, please include the purchase order number or Abcam order reference number so I can locate your order on our system.

Thanks in advance for the additional information.

Merci de nous avoir contactés. Nous sommes désolés d'apprendre que le produit que vous avez reçu ne fonctionne pas comme attendu. Comme convenu j'ai mis en place l'envoi d'une unité de remplacement avec un lot différent, le numéro de commande de remplacement gratuit de ce produit : ******. Vous recevrez prochainement un mail de confirmation comprenant les détails d'expédition. N'hésitez pas à nous contacter lors d'une prochaine occasion.

Thank you for contacting us and I'm sorry to hear that this antibody is giving you some trouble. I think that boiling will more properly denature the protein and allow it to move more efficiently through the gel. To denature, use a loading buffer with SDS and boil the mixture at 95-100°C for 5 minutes. You can find our general WB protocol here: https://www.abcam.com/ps/pdf/protocols/WB-beginner.pdf Hopefully this will improve the resolution of the band, but it probably wouldn't hurt to do a nuclear fractionation to isolate the protein of interest. Our protocol is specific for cells: https://www.abcam.com/index.html?pageconfig=resource&rid=11408 but there is some literature on isolation of nuclei from tissues: http://www.ncbi.nlm.nih.gov/pubmed/18228353 I don't think you'll need it, but I just wanted to make you aware that we do carry a new nuclear extraction kit for cells and tissues that was just added to our catalogue yesterday: https://www.abcam.com/EpiSeeker-Nuclear-Extraction-Kit-ab113474.html I hope this information helps. Please contact us with any other questions.

I'm very sorry to hear you are still having no staining with the recent set of experiments, I appreciate the effort you have put to make the experiments work and cannot understand why the antibody did not work for you. We have not received any other complaints on this product in a year so I suspect your vial was damaged during shipping or storage. I would be happy to offer you a refund, credit note or replacement vial if you can tell me which you would prefer. I look forward to hearing from you to arrange this,

I am pleased to inform you that I have received just this morning a detailed protocol for staining with ab11054. The antibody has been very satisfactory in detecting the Dmc1 protein on mouse meiotic chromosomes. The meiotic chromosomes spreads were performed as described in the protocol that I have added to the online datasheet (accessible via the link below). Which means cells were fixed using a 4% paraformaldehyde, solution and permeabilized using SDS (0.03% in a 4% paraformaldehyde solution). The IF localization of Dmc1 was then conducted as described in the attached protocol. The 2H12/4 antibody was diluted 1/100 In 500µl of 1X ADB and incubated overnight at 4ºC. The secondary antibody used was a Goat anti-Mouse antibody labeled with Alexa-488 dye (A11017, Invitrogen), diluted 1/500. The slides were then mounted in a drop of Vectashield mounting medium with DAPI (H-1500, Vector Laboratories). This procedure allowed localization of the Dmc1 protein as numerous foci in the Leptotene and Zygotene stages of meiotic prophase, and fewer foci in the early Pachytene stage of meiotic prophase. I hope this information will be useful, please let me know if you still experience problems,

I'm very sorry for the delay in finding out more information to help you and to hear you are experiencing problems with two of our antibodies. I have been informed that ab36453 has been used in the following reference: Geddis AE & Kaushansky K. Megakaryocytes express functional Aurora-B kinase in endomitosis. Blood 104:1017-24 (2004). PubMed: 15130946 I enclose the staining protocol for this antibody: ICC/IF: 1/1000. Fix cells in suspension by the addition of an equal volume of 4% paraformaldehyde prepared in phosphate-buffered saline (PBS, pH 7.4) for a final concentration of 2% paraformaldehyde. Fix cells at room temperature for 45 minutes and then centrifuge and wash twice with PBS. Resuspend cells in PBST (PBS containing 0.5% Triton-X100) with 3% bovine serum albumin (BSA) and incubate at room temperature for 15 to 30 minutes to provide blocking against nonspecific antibody reactivity. Unfortunately I am still trying to find out the staining protocol with ab11054, I apologize for the delay. I would like to offer for me to look at your protocol as I may be able to make suggestions to improve the staining. I enclose the link below which will take you to a short questionnaire. If you please put it to my attention I will be able to continue my investigations and hope to help you resolve this problem: https://www.abcam.com/index.html?section=ihc&pageconfig=technical&intAbID=11054&mode=questionaire I look forward to hearing from you and hope the protocol for ab36453 will be useful,

Thank you for your enquiry. I am sorry to hear that your customer has been having difficulties with this antibody. Quality is important to Abcam. Your customer is using a western blotting approach that I would largely recommend. I would be very interested to know whether this cross reacting product is the major product on the blot or whether this is a mild cross reacting band. I would therefore appreciate it if you could ask your customer if they could -mail me a blot image as this would be most informative. We take all complaints very seriously and if there is a major cross reacting band we need to consider to remove it from our catalogue. I look forward to hearing from you.