Anti NOX2  

Immunohistochemistry analysis of NOX2/CYBB using anti-NOX2 antibody. NOX2 was detected in a paraffin-embedded section of human tonsil 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-NOX2 Antibody 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 with DAB as the chromogen.

Western blot analysis of NOX2/CYBB using anti-NOX2 antibody. 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 Jurkat whole cell lysates, Lane 3: human MCF-7 whole cell lysates, Lane 4: rat RH35 whole cell lysates, Lane 5: mouse RAW264.7 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 affinity purified rabbit anti-NOX2 antigen polyclonal antibody 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 with Tanon 5200 system. A specific band was detected for NOX2 at approximately 65 kDa. The expected band size for NOX2 is at 65 kDa.

Western blot analysis analysis of NOX2/gp91phox/CYBB using anti-NOX2/gp91phox/CYBB antibody. 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 30μg of sample under reducing conditions. After electrophoresis, proteins were transferred to a nitrocellulose membrane at 150 mA for 50-90 minutes. Blocked the membrane with 5% milk for 1.5 hour at RT. The membrane was incubated with rabbit anti-NOX2/gp91phox/CYBB antibody at 1:1000 overnight at 4°C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a Cell Signaling Technology's 7074s murine anti-rabbit conformation specific antibody at a dilution of 1:5000 for 1 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit with Tanon 5200 system. A specific band was detected for NOX2/gp91phox/CYBB at approximately 65 kDa. The expected band size for NOX2/gp91phox/CYBB is at 65 kDa.