TBR1 Antibody / T-box brain protein 1 (FY13103)

Flow Cytometry analysis of SH-SY5Y cells using anti-TBR1 antibody. Overlay histogram showing SH-SY5Y cells stained with (Blue line). To facilitate intracellular staining, cells were fixed with 4% paraformaldehyde and permeabilized with permeabilization buffer. The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-TBR1 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 TBR1 using anti-TBR1 antibody. Lane 1: human SH-SY5Y whole cell lysates, Lane 2: rat brain tissue lysates, Lane 3: rat C6 whole cell lysates, Lane 4: mouse brain tissue lysates, Lane 5: mouse Neuro-2a whole cell 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-TBR1 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 enhanced chemiluminescent. TBR1 antibody detects a doublet at ~70-75 kDa across the indicated brain-derived samples and cell lines. TBR1’s calculated mass is ~74 kDa; the closely spaced pair is consistent with differential post-translational modification (commonly phosphorylation) and is frequently observed for this nuclear transcription factor.