IFI30 Antibody / GILT / Gamma-interferon-inducible lysosomal thiol reductase (FY13101)

Flow Cytometry analysis of HepG2 cells using anti-IFI30 antibody. Overlay histogram showing HepG2 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-IFI30 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 (Red line) was also used as a control.

Western blot analysis of IFI30 using anti-IFI30 antibody. Lane 1: human Raji whole cell lysates, Lane 2: human HepG2 whole cell lysates, Lane 3: rat liver tissue lysates, Lane 4: rat spleen tissue lysates, Lane 5: 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-IFI30 antibody at 0.25 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. IFI30 antibody detects a predominant band at ~23 kDa with a minor band at ~30 kDa across the indicated lysates. IFI30 is synthesized as a ~35 kDa glycoprotein and processed in the endo-lysosomal pathway to a mature form typically near ~28-30 kDa; additional C-terminal trimming and glycosylation differences can yield a smaller ~23-24 kDa species. The observed pattern is consistent with mature IFI30 predominating alongside a minor precursor or intermediate.