Anti-Caspase-9/Casp9 Antibody Picoband®

Figure 1. Western blot analysis of Caspase-9/Casp9 using anti-Caspase-9/Casp9 antibody (A00080-7).
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: mouse liver tissue lysates,
Lane 2: mouse pancreas tissue lysates,
Lane 3: mouse stomach 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-Caspase-9/Casp9 antigen affinity purified polyclonal antibody (Catalog # A00080-7) 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 Caspase-9/Casp9 at approximately 46 kDa. The expected band size for Caspase-9/Casp9 is at 46 kDa.

Figure 2. IF analysis of Caspase-9/Casp9 using anti-Caspase-9/Casp9 antibody (A00080-7).
Caspase-9/Casp9 was detected in an immunocytochemical section of HEPA1-6 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-Caspase-9/Casp9 Antibody (A00080-7) 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. 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 mouse spleen tissues using anti-Caspase-9/Casp9 antibody (A00080-7).
Overlay histogram showing mouse spleen tissue stained with A00080-7 (Blue line). To facilitate intracellular staining, tissues were fixed with 4% paraformaldehyde and permeabilized with permeabilization buffer. The tissue sections were blocked with 10% normal goat serum. And then incubated with rabbit anti-Caspase-9/Casp9 Antibody (A00080-7, 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 without incubation with primary antibody and secondary antibody (Red line) was used as a blank control.

Figure 4. IHC analysis of Caspase-9/Casp9 using anti-Caspase-9/Casp9 antibody (A00080-7).
Caspase-9/Casp9 was detected in a paraffin-embedded section of mouse brain 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 2 μg/ml rabbit anti-Caspase-9/Casp9 Antibody (A00080-7) 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.

Figure 5. IHC analysis of Caspase-9/Casp9 using anti-Caspase-9/Casp9 antibody (A00080-7).
Caspase-9/Casp9 was detected in a paraffin-embedded section of rat brain 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 2 μg/ml rabbit anti-Caspase-9/Casp9 Antibody (A00080-7) 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.