Anti-FADD Antibody

• Catalog Number: orb692173
• Category: Antibodies
• Description: Anti-FADD Antibody. Tested in Flow Cytometry, IHC, WB applications. This antibody reacts with Mouse, Rat.
• Clonality: Polyclonal
• Species/Host: Rabbit
• Isotype: Rabbit IgG
• Conjugation: Unconjugated
• Reactivity: Mouse, Rat
• Form/Appearance: Lyophilized
• Concentration: Adding 0.2 ml of distilled water will yield a concentration of 500 μg/ml.
• Immunogen: A synthetic peptide corresponding to a sequence at the C-terminus of mouse FADD, which shares 63.6% and 81.8% amino acid (aa) sequence identity with human and rat FADD, respectively.
• UniProt ID: Q61160
• MW: 30 kDa
• Tested applications: FC, IHC, WB
• Application notes: Western blot, 0.1-0.25μg/ml, Mouse, Rat Immunohistochemistry (Paraffin-embedded Section), 2-5μg/ml, Mouse Flow Cytometry (Fixed), 1-3μg/1x106 cells, Mouse. Add 0.2ml of distilled water will yield a concentration of 500μg/ml
• Antibody Type: Primary Antibody
• Storage: Maintain refrigerated at 2-8°C for up to 2 weeks. For long term storage store at -20°C in small aliquots to prevent freeze-thaw cycles.
• Alternative names: Hemoglobin subunit alpha; Alpha-globin; Hemoglobin
• Note: For research use only

Flow Cytometry analysis of HEPA1-6 cells using anti-FADD antibody. Overlay histogram showing HEPA1-6 cells (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-FADD Antibody (1 µg/1x10^6 cells) for 30 min at 20°C. DyLight®488 conjugated goat anti-rabbit IgG (5-10 µg/1x10^6 cells) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was rabbit IgG (1 µg/1x10^6) used under the same conditions. Unlabelled sample without incubation with primary antibody and secondary antibody (Red line) was used as a blank control.

IHC analysis of FADD using anti-FADD antibody. FADD was detected in a paraffin-embedded section of mouse liver tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 2 µg/ml rabbit anti-FADD Antibody 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) with DAB as the chromogen.

Western blot analysis of FADD using anti-FADD 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: mouse brain tissue lysates, Lane 2: mouse stomach tissue lysates, Lane 3: mouse lung tissue lysates, Lane 4: mouse liver tissue lysates, Lane 5: mouse kidney tissue lysates, Lane 6: mouse spleen tissue lysates, Lane 7: mouse thymus tissue lysates, Lane 8: mouse NIH/3T3 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-FADD antigen affinity purified polyclonal antibody at 0.25 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 FADD at approximately 30 KD. The expected band size for FADD is at 23 KD.

Western blot analysis of FADD using anti-FADD 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: rat brain tissue lysates, Lane 2: rat stomach tissue lysates, Lane 3: rat lung tissue lysates, Lane 4: rat liver tissue lysates, Lane 5: rat kidney tissue lysates, Lane 6: rat spleen tissue lysates, Lane 7: rat thymus tissue lysates, Lane 8: rat PC-12 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-FADD antigen affinity purified polyclonal antibody at 0.25 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 FADD at approximately 30 KD. The expected band size for FADD is at 23 KD.