Figure 1. Western blot analysis of NLRP3 using anti-NLRP3 antibody (A00034-2). 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 30ug of sample under reducing conditions. Lane 1: human Hela whole cell lysates, Lane 2: human U-87MG whole cell lysates, Lane 3: rat spleen tissue lysates, Lane 4: rat PC-12 whole cell lysates, Lane 5: mouse thymus tissue lysates, Lane 6: mouse lung tissue lysates, Lane 7: mouse spleen tissue lysates. 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-NLRP3 antigen affinity purified polyclonal antibody (Catalog # A00034-2) at 0.5 microg/mL 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 1:5000 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 NLRP3 at approximately 118KD. The expected band size for NLRP3 is at 118KD.