Western blot analysis of C3 expression in HepG2 cell lysate (AAA11693).
Electrophoresis was performed on a 5-20% SDS-PAGE gel at 70V (stacking gel) / 90V (resolving gel) for 2-3 hours. The sample well of each lane was loaded with 50ug of sample under reducing conditions. After Electrophoresis, proteins were transferred to a Nitrocellulose membrane at 150mA for 50-90 minutes. Blocked the membrane with 5% Non-fat Milk/TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti-C3 monoclonal antibody (AAA11693) overnight at 4°C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with goat anti-rabbit IgG-HRP secondary antibody at a dilution of 1:10000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit (please inquire) with Tanon 5200 system. A specific band was detected for C3.