ZAP70 Antibody (monoclonal, 9D5)

• Catalog Number: orb654285
• Category: Antibodies
• Description: ZAP70 Antibody (monoclonal, 9D5)
• Species/Host: Mouse
• Clonality: Monoclonal
• Clone Number: 9D5
• Tested applications: FC, WB
• Reactivity: Human, Mouse, Rat
• Isotype: Mouse IgG2b
• Immunogen: A synthetic peptide corresponding to a sequence in the middle region of human ZAP70 (359-393aa MRKKQIDVAIKVLKQGTEKADTEEMMREAQIMHQL), different from the related mouse sequence by two amino acids.
• Concentration: Adding 0.2 ml of distilled water will yield a concentration of 500 μg/ml.
• Dilution range: Western blot, 0.1-0.5μg/ml, Human, Mouse, Rat Flow Cytometry, 1-3μg/1x106 cells, Human
• Form/Appearance: Lyophilized
• Conjugation: Unconjugated
• MW: 72 kDa
• UniProt ID: P43403
• Sensitivity: > 5000 cells
• 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: SRK; STD; TZK; ZAP70; ZAP-70; ZAP 70; P43403
• 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 ZAP70 using anti-ZAP70 antibody (orb654285). 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 Jurkat whole cell lysates; Lane 2: human CCRF-CEM whole cell lysates; Lane 3: rat thymus tissue lysates; Lane 4: mouse spleen tissue lysates; Lane 5: mouse thymus tissue lysates. 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 mouse anti-ZAP70 antigen affinity purified monoclonal antibody (Catalog # orb654285) 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-mouse IgG-HRP secondary antibody at a dilution of 1:10000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit (Catalog # orb90502) with Tanon 5200 system. A specific band was detected for ZAP70 at approximately 72KD. The expected band size for ZAP70 is at 70KD.

Flow Cytometry analysis of 293T cells using anti-ZAP70 antibody (orb654285). Overlay histogram showing 293T cells stained with orb654285 (Blue line). The cells were blocked with 10% normal goat serum. And then incubated with mouse anti-ZAP70 Antibody (orb654285, 1μg/1x10^6 cells) for 30 min at 20°C. DyLight®488 conjugated goat anti-mouse IgG (5-10μg/1x10^6 cells) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was mouse IgG (1μg/1x10^6) used under the same conditions. Unlabelled sample (Red line) was also used as a control.

Flow Cytometry analysis of Jurkat cells using anti-ZAP70 antibody (orb654285). Overlay histogram showing Jurkat cells stained with orb654285 (Blue line). The cells were blocked with 10% normal goat serum. And then incubated with mouse anti-ZAP70 Antibody (orb654285, 1μg/1x10^6 cells) for 30 min at 20°C. DyLight®488 conjugated goat anti-mouse IgG (5-10μg/1x10^6 cells) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was mouse IgG (1μg/1x10^6) used under the same conditions. Unlabelled sample (Red line) was also used as a control.