Phospho-ZAP70/Syk (Tyr319, Tyr352) Monoclonal Antibody (n3kobu5), APC, eBioscience™, Invitrogen™

Mouse Monoclonal Antibody

Supplier:  Invitrogen 17900642

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Catalog No. 50-112-9181

Description

Description: This n3kobu5 monoclonal antibody recognizes human and mouse zeta chain-associated protein of 70 kD (also known as ZAP-70) and spleen tyrosine kinase (also known as SYK) when phosphorylated on Y319 and Y352, respectively. ZAP-70 and SYK are members of the SYK protein tyrosine kinase (PTK) family that are phosphorylated and activated by Src family PTKs. ZAP-70/SYK Y319/Y352 are located in the so-called interdomain of ZAP-70/SYK (between the N-terminal dual SH2 domains and the C-terminal kinase domain). Phosphorylation of ZAP-70 Y319 by Lck is necessary for T cell receptor (TCR)-dependent association of ZAP-70 with Lck and phospholipase C gamma and subsequent activation of calcium-dependent and Ras signaling cascades. SYK Y352 phosphorylation by Fyn/Lyn is critical for propagation of B cell receptor (BCR) signaling and for B cell development. Specificity of this n3kobu5 clone was determined by ELISA, flow cytometry, and western blotting. Applications Reported: This n3kobu5 antibody has been reported for use in intracellular staining followed by flow cytometric analysis. Applications Tested: This n3kobu5 antibody has been pre-titrated and tested by intracellular staining followed by flow cytometric analysis of stimulated normal human peripheral blood cells. This can be used at 5 µL (0.03 µg) per test. A test is defined as the amount (µg) of antibody that will stain a cell sample in a final volume of 100 µL.

Cell number should be determined empirically but can range from 10^5 to 10^8 cells/test. Staining Protocol: All protocols work well for this monoclonal antibody. Use of Protocol A: Two-step protocol: intracellular (cytoplasmic) proteins allows for the greatest flexibility for detection of surface and intracellular (cytoplasmic) proteins. Use of Protocol B: One-step protocol: intracellular (nuclear) proteins is recommended for staining of transcription factors in conjunction with surface and phosphorylated intracellular (cytoplasmic) proteins. Protocol C: Two-step protocol: Fixation/Methanol allows for the greatest discrimination of phospho-specific signaling between unstimulated and stimulated samples, but with limitations on the ability to stain specific surface proteins (refer to "Clone Performance Following Fixation/Permeabilization" located in the Best Protocols Section under the Resources tab online). All Protocols can be found in the Flow Cytometry Protocols: "Staining Intracellular Antigens for Flow Cytometry Protocol" located in the Best Protocols Section under the Resources tab online. Excitation: 633-647 nm; Emission: 660 nm; Laser: Red Laser. Filtration: 0.2 μm post-manufacturing filtered. ZAP-70 and SYK are members of the SYK protein tyrosine kinase (PTK) family that are phosphorylated and activated by Src family PTKs. ZAP-70/SYK Y319/Y352 are located in the so-called interdomain of ZAP-70/SYK (between the N-terminal dual SH2 domains and the C-terminal kinase domain). Phosphorylation of ZAP-70 Y319 by Lck is necessary for T cell receptor (TCR)-dependent association of ZAP-70 with Lck and phospholipase C gamma and subsequent activation of calcium-dependent and Ras signaling cascades. SYK Y352 phosphorylation by Fyn/Lyn is critical for propagation of B cell receptor (BCR) signaling and for B cell development.

Specifications

• Antigen: Phospho-ZAP70/Syk (Tyr319, Tyr352)
• Classification: Monoclonal
• Concentration: 5 μL/Test
• Formulation: PBS with 0.1% gelatin, 0.2% BSA and 0.09% sodium azide; pH 7.2
• Gene Accession No.: P43404, P48025, P43405, P43403
• Host Species: Mouse
• Quantity: 100 Tests
• Primary or Secondary: Primary
• Target Species: Human, Mouse
• Product Type: Antibody
• Isotype: IgG2b κ

• Applications: Flow Cytometry
• Clone: n3kobu5
• Conjugate: APC
• Gene: SYK
• Gene Symbols: Syk, ZAP70
• Purification Method: Affinity chromatography
• Regulatory Status: RUO
• Gene ID (Entrez): 20963, 22637, 6850, 7535
• Content And Storage: 4° C, store in dark, DO NOT FREEZE!
• Form: Liquid

For Research Use Only.