Anti-DKK3/RIG Antibody Picoband™

Figure 1. Western blot analysis of DKK3/RIG using anti-DKK3/RIG antibody (A03140).
Electrophoresis was performed on a 5-20% SDS-PAGE gel at 70V (Stacking gel) / 90V (Resolving gel) for 2-3 hours. The sample well of each lane was loaded with 30 ug of sample under reducing conditions.
Lane 1: human SH-SY5Y whole cell lysates,
Lane 2: human U251 whole cell lysates,
Lane 3: human U20S whole cell lysates,
Lane 4: human U-87MG whole cell lysates,
Lane 5: rat brain tissue lysates,
Lane 6: rat C6 whole cell lysates,
Lane 7: mouse brain tissue lysates,
Lane 8: 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-DKK3/RIG antigen affinity purified polyclonal antibody (Catalog # A03140) 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 Enhanced Chemiluminescent detection (ECL) kit (Catalog # EK1002) with Tanon 5200 system. A specific band was detected for DKK3/RIG at approximately 22 kDa. The expected band size for DKK3/RIG is at 12 kDa.

Figure 2. IF analysis of DKK3/RIG using anti-DKK3/RIG antibody (A03140).
DKK3/RIG was detected in an immunocytochemical section of HELA 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-DKK3/RIG Antibody (A03140) overnight at 4°C. Cy3 Conjugated Goat Anti-Rabbit IgG (BA1032) was 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.

Figure 3. Flow Cytometry analysis of SH-SY5Y cells using anti-DKK3/RIG antibody (A03140).
Overlay histogram showing SH-SY5Y cells stained with A03140 (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-DKK3/RIG Antibody (A03140, 1 μg/1x10⁶ cells) for 30 min at 20°C. DyLight®488 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 (Red line) was also used as a control.