Western blot analysis of Caspase-1 using anti-Caspase-1 antibody (M00048-2).
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 THP-1 whole cell lysates,
Lane 2: human SiHa whole cell lysates,
Lane 3: human A431 whole cell lysates,
Lane 4: human RT4 whole cell lysates,
Lane 5: mouse EL-4 whole cell lysates,
Lane 6: mouse J774A.1 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-Caspase-1 antigen affinity purified monoclonal antibody (Catalog # M00048-2) at 1:500 overnight at 4°C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody at a dilution of 1:500 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit (Catalog # EK1002) with Tanon 5200 system. A specific band was detected for Caspase-1 at approximately 45 kDa. The expected band size for Caspase-1 is at 45 kDa.