Caspase-3 Antibody from (CST)

Results Summary

This antibody is quite hard to optimize, but a dilution of 1:1000 overnight can lead to some good result, depending on the western blot and cell lysate conditions. Hela cells WCL has been loaded on a 4-12% SDS-precast gel (1:15h; 180 V) then proteins have been transferred on nitrocellulose membrane using the semi-dry blotting (1h 11V). Blocking with milk 5% in PBS 0,05% tween (1 hour) was followed by incubation overnight in a cold room with Caspase-3 antibody (1:1000 in milk1% in PBS 0,05% tween) Then the membrane was washed with 3x 5 minutes with PBS 0,05% tween, followed by a 1-hour incubation with Rabbit secondary antibody 1:3000 in milk 1% in PBS 0,05% tween. To wash out the secondary antibody, a second wash with PBS 0,05% tween (3x 5 minutes) was performed. ECL substrate has been used to detect the signal.