Western blot analysis of beta-Actin using anti-beta-Actin antibody (A01263-HRP).
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 30 ug of sample under reducing conditions.
Lane 1: human Hela whole cell lysates,
Lane 2: human HepG2 whole cell lysates,
Lane 3: human MCF-7 whole cell lysates,
Lane 4: human K562 whole cell lysates,
Lane 5: human A549 whole cell lysates,
Lane 6: human A431 whole cell lysates,
Lane 7: human U87 whole cell lysates,
Lane 8: monkey COS-7 whole cell lysates.
After electrophoresis, proteins were transferred to a nitrocellulose membrane at 150 mA 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-beta-Actin antigen affinity purified polyclonal antibody (Catalog # A01263-HRP) at 0.1 μg/mL overnight at 4°C, then washed with TBS-0.1%Tween 3 times with 5 minutes each. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit (Catalog # EK1002) with Tanon 5200 system. A specific band was detected for beta-Actin at approximately 42 kDa. The expected band size for beta-Actin is at 42 kDa.