Anti-JAB1/COPS5 Antibody Picoband®

Western blot analysis of JAB1 using anti-JAB1 antibody (PB9490).
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 Raji whole cell lysates,
Lane 2: human K562 whole cell lysates,
Lane 3: human PC-3 whole cell lysates,
Lane 4: human Hela whole cell lysates,
Lane 5: human A549 whole cell lysates,
Lane 6: human HEK293 whole cell lysates,
Lane 7: rat brain tissue lysates,
Lane 8: rat heart tissue lysates,
Lane 9: mouse brain tissue lysates,
Lane 10: mouse heart tissue lysates,
Lane 11: mouse SP2/0 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-JAB1 antigen affinity purified polyclonal antibody (Catalog # PB9490) 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 JAB1 at approximately 38 kDa. The expected band size for JAB1 is at 38 kDa.

IHC analysis of JAB1 using anti-JAB1 antibody (PB9490).
JAB1 was detected in a paraffin-embedded section of mouse testis tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1 μg/ml rabbit anti-JAB1 Antibody (PB9490) 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 JAB1 using anti-JAB1 antibody (PB9490).
JAB1 was detected in a paraffin-embedded section of rat testis tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1 μg/ml rabbit anti-JAB1 Antibody (PB9490) 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 JAB1 using anti-JAB1 antibody (PB9490).
JAB1 was detected in a paraffin-embedded section of human mammary cancer tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1 μg/ml rabbit anti-JAB1 Antibody (PB9490) 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.

Flow Cytometry analysis of U20S cells using anti-JAB1 antibody (PB9490).
Overlay histogram showing U20S cells stained with PB9490 (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-JAB1 Antibody (PB9490,1μg/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 without incubation with primary antibody and secondary antibody (Red line) was used as a blank control.

Flow Cytometry analysis of U937 cells using anti-JAB1 antibody (PB9490).
Overlay histogram showing U937 cells stained with PB9490 (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-JAB1 Antibody (PB9490,1μg/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 without incubation with primary antibody and secondary antibody (Red line) was used as a blank control.

ICC analysis of JAB1 using anti-JAB1 antibody (PB9490).
JAB1 was detected in immunocytochemical section of A431 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 1μg/ml rabbit anti-JAB1 Antibody (PB9490) overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The section was developed using Strepavidin-Biotin-Complex (SABC)(Catalog # SA1022) with DAB as the chromogen.

Immunoprecipitating (IP) JAB1 in A549 whole cell lysate.
Western blot analysis of JAB1 using anti-JAB1 antibody (PB9490);
Lane 1: A549 whole cell lysates (30ug);
Lane 2: Rabbit control IgG instead of anti-JAB1 antibody in A549 whole cell lysate;
Lane 3: anti-JAB1 antibody (2μg) + A549 whole cell lysate (500μg).
After electrophoresis, proteins were transferred to a membrane. Then the membrane was incubated with rabbit anti-JAB1 antigen affinity purified polyclonal antibody (PB9490) 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 JAB1 at approximately 38 kDa. The expected band size for JAB1 is at 38 kDa.