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Hi, I've attached two images of a fluorescent stain I've done using your SOX2 antibody (part of the mouse embryonic stem cell marker panel- ab107156). The tissue is mouse brain tissue, fixed in 4% paraformaldehyde, sliced with a cryostat at 20 microns thick. I used our most basic fluorescent protocol to start (incubated in the primary antibody at 1:1000 overnight at 4 degrees and then secondary at 1:200 for 2 hours at room temperature). I'm wondering if, in your opinion, the stain worked? Is this what it should look like? Can you suggest any protocol changes to improve it?
Asked on Aug 31 2012
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Based on other images for this product and other SOX2 antibodies, this staining pattern does look correct. However, it seems that there is room for improvement with this staining. Do you permeablize your tissues? Do you block in serum? I did see that one of the Abreviews for this product achieved success using glycine blocking. That same review also used an antigen retrieval, which while uncommon in IHC-Fr, can help with any paraformaldehyde fixation. Have you had the opportunity to run a series dilution with the antibody to determine the best concentration in this protocol? Do your washes contain a mild detergent? I would be very happy to review the protocol which you have used and offer suggestions if you provide that to me.
I hope this information is helpful to you and look forward to your reply. Please do not hesitate to contact us if you need any more advice or information.
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Answered on Aug 31 2012