Anti-p47 phox/NCF1 Antibody Picoband®

Figure 1. Western blot analysis of P47 Phox/NCF1 using anti-P47 Phox/NCF1 antibody (A01586-3).
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 THP-1 whole cell lysates,
Lane 3: human Ramos whole cell lysates,
Lane 4: rat spleen tissue lysates,
Lane 5: mouse thymus tissue lysates,
Lane 6: mouse spleen 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-P47 Phox/NCF1 antigen affinity purified polyclonal antibody (Catalog # A01586-3) 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 P47 Phox/NCF1 at approximately 40-50 kDa. The expected band size for P47 Phox/NCF1 is at 45 kDa.

Figure 2. IHC analysis of P47 Phox/NCF1 using anti-P47 Phox/NCF1 antibody (A01586-3).
P47 Phox/NCF1 was detected in a paraffin-embedded section of rat lymph 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-P47 Phox/NCF1 Antibody (A01586-3) overnight at 4°C. Peroxidase Conjugated Goat Anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using HRP Conjugated Rabbit IgG Super Vision Assay Kit (Catalog # SV0002) with DAB as the chromogen.

Figure 3. IHC analysis of P47 Phox/NCF1 using anti-P47 Phox/NCF1 antibody (A01586-3).
P47 Phox/NCF1 was detected in a paraffin-embedded section of mouse lymph 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-P47 Phox/NCF1 Antibody (A01586-3) overnight at 4°C. Peroxidase Conjugated Goat Anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using HRP Conjugated Rabbit IgG Super Vision Assay Kit (Catalog # SV0002) with DAB as the chromogen.

Figure 4. Flow Cytometry analysis of THP-1 cells using anti-P47 Phox/NCF1 antibody (A01586-3).
Overlay histogram showing THP-1 cells stained with A01586-3 (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-P47 Phox/NCF1 Antibody (A01586-3, 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.

Figure 5. IF analysis of Phox/NCF1 using anti-Phox/NCF1 antibody (A01586-3).
Phox/NCF1 was detected in a paraffin-embedded section of mouse lymph node 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 5 μg/mL rabbit anti-Phox/NCF1 Antibody (A01586-3) overnight at 4°C. DyLight®594 Conjugated Goat Anti-Rabbit IgG (BA1142) 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 6. IF analysis of Phox/NCF1 using anti-Phox/NCF1 antibody (A01586-3).
Phox/NCF1 was detected in a paraffin-embedded section of rat lymph node 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 5 μg/mL rabbit anti-Phox/NCF1 Antibody (A01586-3) overnight at 4°C. DyLight®594 Conjugated Goat Anti-Rabbit IgG (BA1142) 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.