ATG7 Antibody

• Catalog Number: orb654295
• Category: Antibodies
• Description: ATG7 Antibody
• Species/Host: Rabbit
• Clonality: Polyclonal
• Tested applications: FC, IHC, WB
• Reactivity: Human, Mouse, Rat
• Isotype: Rabbit IgG
• Immunogen: A synthetic peptide corresponding to a sequence of human ATG7 (HELTQKKLNEYRLDEAPKDIK).
• Concentration: Adding 0.2 ml of distilled water will yield a concentration of 500 μg/ml.
• Dilution range: Western blot, 0.1-0.25μg/ml, Human, Mouse, Rat Immunohistochemistry (Paraffin-embedded Section), 0.5-1μg/ml, Human, Mouse Flow Cytometry, 1-3μg/1x106 cells, Human, Mouse, Rat
• Form/Appearance: Lyophilized
• Conjugation: Unconjugated
• MW: 78 kDa
• UniProt ID: O95352
• Storage: Store at -20˚C for one year from date of receipt. After reconstitution, at 4˚C for one month. It can also be aliquotted and stored frozen at -20˚C for six months. Avoid repeated freeze-thaw cycles.
• Alternative names: Ubiquitin-like modifier-activating enzyme ATG7; AT
• Note: For research use only
• Application notes: Tested Species: In-house tested species with positive results. Other applications have not been tested. Optimal dilutions should be determined by end users. Add 0.2ml of distilled water will yield a concentration of 500ug/ml.
• Expiration Date: 12 months from date of receipt.

Western blot analysis of ATG7 using anti-ATG7 antibody (orb654295). 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 50ug of sample under reducing conditions. Lane 1: human CACO-2 whole cell lysates, Lane 2: human U20S whole cell lysates, Lane 3: human HEPG2 whole cell lysates, Lane 4: human HEK293 whole cell lysates, Lane 5: human HELA whole cell lysates, Lane 6: human K562 whole cell lysates, Lane 7: rat thymus tissue lysates, Lane 8: rat spleen tissue lysates, Lane 9: mouse SP2/0 whole cell lystates. After Electrophoresis, proteins were transferred to a Nitrocellulose membrane at 150mA 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-ATG7 antigen affinity purified polyclonal antibody (Catalog # orb654295) 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 (Catalog # orb90503) with Tanon 5200 system. A specific band was detected for ATG7 at approximately 78KD. The expected band size for ATG7 is at 78KD.

Flow Cytometry analysis of C6 cells using anti-ATG7 antibody (orb654295). Overlay histogram showing C6 cells stained with orb654295 (Blue line). The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-ATG7 Antibody (orb654295, 1μg/1x10^6 cells) for 30 min at 20°C. DyLight®488 conjugated goat anti-rabbit IgG (5-10μg/1x10^6 cells) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was rabbit IgG (1μg/1x10^6) used under the same conditions. Unlabelled sample (Red line) was also used as a control.

Flow Cytometry analysis of HELA cells using anti-ATG7 antibody (orb654295). Overlay histogram showing HELA cells stained with orb654295 (Blue line). The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-ATG7 Antibody (orb654295, 1μg/1x10^6 cells) for 30 min at 20°C. DyLight®488 conjugated goat anti-rabbit IgG (5-10μg/1x10^6 cells) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was rabbit IgG (1μg/1x10^6) used under the same conditions. Unlabelled sample (Red line) was also used as a control.

Flow Cytometry analysis of RAW264.7 cells using anti-ATG7 antibody (orb654295). Overlay histogram showing RAW264.7 cells stained with orb654295 (Blue line). The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-ATG7 Antibody (orb654295, 1μg/1x10^6 cells) for 30 min at 20°C. DyLight®488 conjugated goat anti-rabbit IgG (5-10μg/1x10^6 cells) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was rabbit IgG (1μg/1x10^6) used under the same conditions. Unlabelled sample (Red line) was also used as a control.

IHC analysis of ATG7 using anti-ATG7 antibody (orb654295). ATG7 was detected in paraffin-embedded section of mouse kidney 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 1μg/ml rabbit anti-ATG7 Antibody (orb654295) 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 # orb90444) with DAB as the chromogen.

IHC analysis of ATG7 using anti-ATG7 antibody (orb654295). ATG7 was detected in paraffin-embedded section of human lung cancer 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 1μg/ml rabbit anti-ATG7 Antibody (orb654295) 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 # orb90444) with DAB as the chromogen.