Anti-CD10抗体[56C6] (ab951)

The IgG concentration of ab951 is 7.2µg/mL.

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Thank you for taking time to complete our questionnaire and for contacting us. I am sorry to hear this antibody is not providing satisfactory results.

I am sorry that these protocol details did not yield the expected 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.

I apologize for the inconvenience. If you have contacted us within six months of purchase I am pleased to offer you a free of charge replacement, or refund in compensation. In order to process this could you please provide me with the Abcam order confirmation number or the PO number and date of purchase?

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The concentration of the Anti-CD10 antibody [56C6] (ab951), lot number GR38260-2 is 7.2µg/mL.

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I am sorry that this product has not been performing as it should in mouse.This product is covered by our Abpromise guarantee. We are happy provide scientific support at any time. If you are using the product in species and 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.

If you could provide me with the details of your protocol, I would be very happy to review those and see if there are any suggestions I acn make which will help.

I have also found the protocol used byDevi L et al.Mechanisms underlying insulin deficiency-induced acceleration of ß-amyloidosis in a mouse model of Alzheimer's disease.PLoS One7:e32792 (2012).http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=22403710&dopt=Abstract

They were able to use this product successfully in mouse tissue and their protocol information may help. I have pasted this below:

Hemibrain samples were taken from the mice under deep isoflurane anesthesia and were snap-frozen for biochemical assays. For western blot analysis, each sample was homogenized in 5 volumes of modified RIPA buffer containing 150 mM NaCl, 50 mM Tris HCl (pH 8.0), 1 mM EDTA, 1% IGEPAL, 0.5% sodium deoxycholate, 0.1% SDS and protease/phosphatase inhibitor cocktail (Calbiochem, La Jolla, CA, USA), and centrifuged at 10,000 g for 10 min to remove any insoluble material. Protein concentrations were determined by a BCA protein assay kit (Pierce, Rockford, IL, USA), and 20–50 µg of protein was run on 4–12% NuPAGE gels (Invitrogen, Carlsbad, CA, USA) and transferred to nitrocellulose membranes. After blocking, membranes were probed with anti-neprilysin 1:2,000, (ab951, Abcam, Cambridge, MA, USA). They were then incubated with horseradish peroxidase-conjugated secondary IgG. Immunoblot signals were visualized by an ECL chemiluminescence substrate reagent kit and were quantified by densitometric scanning and image analysis using Quantity One software.

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The IgG concentration for lot GR79429-1 is 7.2µg/mL. For IHC, the antibody should be used at a concentration of 1/20 - 1/50. The antibody was validated for IHC with a sensitive polymer detection system, so if you are using a different kind of detection, for instance an ABC kit, or a conjugated secondary without any ABC or polymer amplification, we recommend the lower dilution of 1/20. A bottle of the diluent Renoir Red is included as the optimal diluent. Just substitute it in place of your standard buffer diluent.

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Thank you for contacting Abcam.

I think you are correct that the band that you are seeing at 100kDa is correct as based on the Swissprot sequence http://www.uniprot.org/uniprot/Q61391, there are q few glycosylation sites on this protein that would account for the increase in the predicted molecular weight. The 75kDa mat be a non-specific band in which case changing your blocking reagent to 5%BSA may help to reduce that signal. Otherwise if it is a breakdown product then you could increase the amount of protease inhibitors that you are using.

The antibody is covered under our Abpromise for six months and is guaranteed to work in western blot on mouse 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.

Please let me know if there is anything else I can help you with.