Anti-DDX18 Antibody Picoband®

Western blot analysis of DDX18 using anti-DDX18 antibody (A10977-1).
Electrophoresis was performed on a 10% SDS-PAGE gel at 80V (Stacking gel) / 120V (Resolving gel) for 2 hours. The sample well of each lane was loaded with 30 ug of sample under reducing conditions.
Lane 1: human HEL whole cell lysates,
Lane 2: human SH-SY5Y whole cell lysates,
Lane 3: human HepG2 whole cell lysates,
Lane 4: human Caco-2 whole cell lysates,
Lane 5: rat C6 whole cell lysates,
Lane 6: rat RH35 whole cell lysates,
Lane 7: mouse Neuro-2a whole cell lysates,
Lane 8: mouse HEPA1-6 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-DDX18 antigen affinity purified polyclonal antibody (A10977-1) at 0.5 μg/mL overnight at 4°C, 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 is developed using an ECL Plus Western Blotting Substrate (Catalog # AR1196-200) with Tanon 5200 system. A specific band was detected for DDX18 at approximately 80 kDa. The expected band size for DDX18 is at 75 kDa.

IF analysis of DDX18 using anti-DDX18 antibody (A10977-1) and anti-Alpha Tubulin antibody (M03989-3).
DDX18 was detected in an immunocytochemical section of U2OS cells. Enzyme antigen retrieval was performed using IHC enzyme antigen retrieval reagent (AR0022) for 15 mins. The cells were blocked with 10% goat serum. And then incubated with 5 μg/mL rabbit anti-DDX18 Antibody (A10977-1) and mouse anti-Alpha Tubulin antibody (M03989-3) overnight at 4°C. Fluoro488 Conjugated Goat Anti-Rabbit IgG (BA1127) and Cy3 Conjugated Goat Anti-Mouse IgG (BA1031) were used as secondary antibody at 1:500 dilution and incubated for 30 minutes at 37°C. The section was counterstained with DAPI. Visualize using a fluorescence microscope and filter sets appropriate for the label used.

Immunoprecipitating DDX18 in HepG2 whole cell lysate.
Western blot analysis of DDX18 using anti-DDX18 antibody (A10977-1).
Lane 1: HepG2 whole cell lysates (30ug),
Lane 2: Rabbit control IgG instead of anti-DDX18 antibody in HepG2 whole cell lysate,
Lane 3: anti-DDX18 antibody (2μg) + HepG2 whole cell lysate (500μg).
After electrophoresis, proteins were transferred to a membrane. Then the membrane was incubated with rabbit anti-DDX18 antigen affinity purified polyclonal antibody (A10977-1) at a dilution of 0.5 μg/mL and probed with a goat anti-rabbit IgG-HRP secondary antibody (Catalog # BA1054). The signal is developed using ECL Plus Western Blotting Substrate (Catalog # AR1197). A specific band was detected for DDX18 at approximately 80 kDa. The expected band size for DDX18 is at 75 kDa.

Flow Cytometry analysis of SH-SY5Y cells using anti-DDX18 antibody (A10977-1).
Overlay histogram showing SH-SY5Y cells stained with A10977-1 (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-DDX18 Antibody (A10977-1, 1 μg/1x10⁶ cells) for 30 min at 20°C. Fluoro488 conjugated goat anti-rabbit IgG (BA1127, 5-10 μg/1x10⁶ cells) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was rabbit IgG (1 μg/1x10⁶) used under the same conditions. Unlabelled sample without incubation with primary antibody and secondary antibody (Red line) was used as a blank control.