Western blot analysis of Cystatin B in Lane 1: human U-87MG whole cell lysate, Lane 2: human SW620 whole cell lysate, Lane 3: human U-937 whole cell lysate, Lane 4: human Caco-2 whole cell lysate using 50 µg (reducing conditions) per well. Electrophoresis was performed on 5-20% SDS-PAGE gel at 70V (Stacking gel) / 90V (Resolving gel) for 2-3 hours and protein was transferred to a nitrocellulose membrane at 150mA for 50-90 minutes. Sample was blocked with 5% Non-fat Milk/TBS for 1.5 hours at room temperature, incubated with Cystatin B monoclonal antibody (Product # MA5-27873) at a dilution of 0.5 µg/mL (overnight at 4°C), followed by goat anti-mouse IgG-Biotin secondary antibody at a dilution of 1:10,000. Signal development was performed using a chemiluminescence (ECL) kit.