Figure 1. Western blot analysis of CYLD using anti-CYLD antibody (PB9543). Electrophoresis was performed on a 5-20% SDS-PAGE gel at 70V (Stacking gel) / 90V (Resolving gel) for 2-3 hours. Lane 1: Rat Testis Tissue Lysate at 50ug, Lane 2: Rat Brain Tissue Lysate at 50ug, Lane 3: HELA Whole Cell Lysate at 40ug. 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-CYLD antigen affinity purified polyclonal antibody (Catalog # PB9543) 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 CYLD at approximately 107 kDa. The expected band size for CYLD is at 107 kDa.