Western blot analysis of Me31B using anti-Fruit fly Me31B antibody (DZ33938-1).
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.
After electrophoresis, proteins were transferred to a nitrocellulose membrane at 150 mA for 50-90 minutes. Blocked the membrane with non-fat dry milk for 1.5 hour at RT. The membrane was incubated with rabbit anti-Fruit fly Me31B antibody (DZ33938-1) at 1:10,000 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 1:10,000, incubation 1h 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 Me31B at approximately 52 kDa. The expected band size for Me31B is at 52 kDa.