Anti-RUNX1T1/ETO Antibody Picoband®

Western blot analysis of RUNX1T1 using anti-RUNX1T1 antibody (A01792-1).
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 Hela whole cell lysates,
Lane 2: human SH-SY5Y 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-RUNX1T1 antigen affinity purified polyclonal antibody (A01792-1) 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 (Catalog # BA1054) 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 RUNX1T1 at approximately 70-80 kDa. The expected band size for RUNX1T1 is at 67 kDa.

IHC analysis of RUNX1T1/ETO using anti-RUNX1T1/ETO antibody (A01792-1).
RUNX1T1/ETO was detected in paraffin-embedded section of human mammary cancer tissue. Heat mediated antigen retrieval was performed in citrate buffer (pH6, epitope retrieval solution) for 20 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1μg/ml rabbit anti-RUNX1T1/ETO Antibody (A01792-1) overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC)(Catalog # SA1022) with DAB as the chromogen.

IHC analysis of RUNX1T1/ETO using anti-RUNX1T1/ETO antibody (A01792-1).
RUNX1T1/ETO was detected in paraffin-embedded section of mouse brain tissue . Heat mediated antigen retrieval was performed in citrate buffer (pH6, epitope retrieval solution) for 20 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1μg/ml rabbit anti-RUNX1T1/ETO Antibody (A01792-1) overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC)(Catalog # SA1022) with DAB as the chromogen.

IF analysis of RUNX1T1 using anti-RUNX1T1 antibody (A01792-1).
RUNX1T1 was detected in immunocytochemical section of U20S cell. 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 2μg/mL rabbit anti-RUNX1T1 Antibody (A01792-1) overnight at 4°C. DyLight®488 Conjugated Goat Anti-Rabbit IgG (BA1127) was used as secondary antibody at 1:100 dilution and incubated for 30 minutes at 37°C. Visualize using a fluorescence microscope and filter sets appropriate for the label used.

Immunoprecipitating (IP) RUNX1T1 in SH-SY5Y whole cell lysate.
Western blot analysis of RUNX1T1 using anti-RUNX1T1 antibody (A01792-1);
Lane 1: SH-SY5Y whole cell lysates (30ug);
Lane 2: Rabbit control IgG instead of anti-RUNX1T1 antibody in SH-SY5Y whole cell lysate;
Lane 3: anti-RUNX1T1 antibody (2μg) + SH-SY5Y whole cell lysate (500μg).
After electrophoresis, proteins were transferred to a membrane. Then the membrane was incubated with rabbit anti-RUNX1T1 antigen affinity purified polyclonal antibody (A01792-1) 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 # AR1196-200). A specific band was detected for RUNX1T1 at approximately 70-80 kDa. The expected band size for RUNX1T1 is at 67 kDa.

Flow Cytometry analysis of U20S cells using anti-RUNX1T1 antibody (A01792-1).
Overlay histogram showing U20S cells stained with A01792-1 (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-RUNX1T1 Antibody (A01792-1,1μg/1x10⁶ cells) for 30 min at 20°C. DyLight®488 conjugated goat anti-rabbit IgG (BA1127, 5-10μg/1x10⁶ cells) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was rabbit IgG (1μg/1x10⁶) used under the same conditions. Unlabelled sample (Red line) was also used as a control.