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Amount of protein loaded: 20 mcg Blocking conditions: 1 hour in 5% NFMP TBST Detection method: Femto ECL and film Electrophoresis and transfer conditions: 12.5% acrylamide gel, 60 minute 15 V semidry transfer Primary antibody conditions (diluent/dilution, incubation time/temperature): 1 mcg/mL in 2.5% NFMP TBST at 4C overnight Secondary antibody conditions (diluent/dilution, incubation time/temperature): 1:2000 goat anti rabbit HRP in 2.5% NFMP TBST 1 hour at room temperature Wash steps: 5 minutes in TBST 3 times, 5 minutes in TBS 2 times prior to developing Primary band is detected around 80-90 kDa instead of the expected 131 kDa. Further development reveals significant secondary bands. Protein was isolated from 5637 human bladder cancer cell line lysed in RIPA with protease inhibitors.
Asked on Oct 31 2012
Thank you for contacting Abcam.
I am sorry that you are having some issues with ab1440. The antibody is covered under our Abpromise for six months and is guaranteed to work in western blot on 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 able to try and aid you further in this matter, would you be able to provide an image of the results that you are recieving, when you use this antibody?
I look forward to your reply and helping to resolve this issue.
Answered on Oct 31 2012