FMOD Antibody / Fibromodulin (FY12023)

Flow Cytometry analysis of Caco-2 cells using anti-FMOD antibody. Overlay histogram showing Caco-2 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-FMOD 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 FMOD using anti-FMOD antibody. Electrophoresis was performed on a 10% SDS-PAGE gel at 80V (Stacking gel) / 120V (Resolving gel) for 2 hours. Lane 1: human HepG2 whole cell lysates, Lane 2: human Hela whole cell lysates, Lane 3: human whole cell lysates, Lane 4: 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-FMOD 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. Expected size of FMOD ~43 kDa (core protein). Observed bands at ~70 kDa (human) and ~60 kDa (mouse) reflect the mature, glycosylated/keratan-sulfate-modified extracellular form of fibromodulin, consistent with published Western blot data showing 70-110 kDa for FMOD in human tissues.