DTNB Antibody / Dystrobrevin beta (FY12628)

Flow Cytometry analysis of Jurkat cells using anti-DTNB antibody. Overlay histogram showing Jurkat cells stained with (Blue line). The cells were fixed with 4% paraformaldehyde and blocked with 10% normal goat serum. And then incubated with rabbit anti-DTNB antibody (1 ug/million cells) for 30 min at 20oC. DyLight 488 conjugated goat anti-rabbit IgG (5-10 ug/million cells) was used as secondary antibody for 30 minutes at 20oC. Isotype control antibody (Green line) was rabbit IgG (1 ug/million cells) used under the same conditions. Unlabelled sample without incubation with primary antibody and secondary antibody (Red line) was used as a blank control.

Western blot analysis of DTNB using anti-DTNB antibody. Electrophoresis was performed on a 10% SDS-PAGE gel at 80V (Stacking gel) / 120V (Resolving gel) for 2 hours. Lane 1: human SH-SY5Y whole cell lysates, Lane 2: human Jurkat whole cell lysates, Lane 3: human 293T whole cell lysates, Lane 4: human SIHA whole cell lysates, Lane 5: rat brain tissue lysates, Lane 6: rat liver tissue lysates, Lane 7: mouse brain tissue lysates, Lane 8: mouse liver tissue lysates. 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-DTNB antibody at 0.5 ug/ml overnight at 4oC, 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 was developed using an ECL Plus Western Blotting Substrate. Western blot probed with anti-DTNB shows a doublet near ~70 kDa in human samples representing distinct phosphorylated or alternatively spliced isoforms of dystrobrevin beta, while mouse and rat samples display a single band corresponding to the conserved major isoform.