Figure 1. Western blot analysis of BubR1/BUB1B using anti-BubR1/BUB1B antibody (M01564-2). 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 50ug of sample under reducing conditions. Lane 1: human Hela whole cell lysates; Lane 2: human HEK293 whole cell lysates; Lane 3: human K562 whole cell 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 mouse anti-BubR1/BUB1B antigen affinity purified monoclonal antibody (Catalog # M01564-2) 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-mouse 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 (Catalog # EK1001) with Tanon 5200 system. A specific band was detected for BubR1/BUB1B at approximately 130KD. The expected band size for BubR1/BUB1B is at 120KD.