RAB34 Antibody / Ras-related protein Rab-34 (FY12985)

Flow Cytometry analysis of cells using anti-RAB34 antibody. Overlay histogram showing 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-RAB34 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 RAB34 using anti-RAB34 antibody. Lane 1: human Hela whole cell lysates, Lane 2: human whole cell lysates, Lane 3: human U2OS whole cell lysates, Lane 4: human whole cell lysates, Lane 5: rat ovary tissue lysates, Lane 6: rat PC-12 whole cell lysates, Lane 7: mouse ovary tissue lysates, Lane 8: mouse 3T3-L1 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-RAB34 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. A single band is detected at ~34 kDa, running above the ~29 kDa prediction. The higher apparent MW is characteristic of lipid-modified Rab GTPases; C-terminal geranylgeranylation (± palmitoylation) alters SDS–PAGE mobility without large mass increase, yielding an observed size near 34 kDa.