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BATCH NUMBER 87891 ORDER NUMBER 26148 DESCRIPTION OF THE PROBLEM I have tried this antibody at 4 different concentrations ranging from 1:250 to 1:2000 and cannot see any staining by immunofluorescence. My staining protocol works for the Invitrogen monoclonal V5 and we have generated an antibody against the protein to which it is tagged and can see the distribution and increased expression by IF and western, respectively, but I see no staining with this polyclonal. If you have defined conditions for this antibody to work, please let me know. Thank you. SAMPLE primary epithelial cells PRIMARY ANTIBODY 1:250, 1:500, 1:1000, 1:2000 4oC overnight, three washes in TBS, blocked with BSA and 5%milk. POSITIVE AND NEGATIVE CONTROLS USED antibody against HA tagged protein, antibody against endogenous protein, Invitrogen monoclonal. SAMPLE PREPARATION 4% paraformaldehyde fixation and 0.5% triton permeabilization. Antibody added overnight at 4C. HOW MANY TIMES HAVE YOU TRIED THE APPLICATION? 4 HAVE YOU RUN A "NO PRIMARY" CONTROL? Yes
Asked on Mar 18 2005
Thank you for your patience. I have heard back from the originator of this antibody and your protocol looks fine, especially since the monoclonal worked under the same conditions. There may have been a problem with the vial/lot that you received and I can certainly offer you a different batch to try, an alternative antibody (I know that you mentioned the chicken polyclonal), or a refund. Please let me know which you would prefer. Have a great weekend.
回复于 Mar 25 2005