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7779
HTScan® Syk Kinase Assay Kit

HTScan® Syk Kinase Assay Kit #7779

This product is discontinued

Kinase Assay - Radiometric Image 1

Figure 1. Syk kinase activity was measured in a radiometric assay using the following reaction conditions: 5 mM MOPS, pH 7.2, 2.5 mM β-glycerophosphate, 1 mM EGTA, 0.4 mM EDTA, 4 mM MgCl2, 2.5 mM MnCl2, 0.05 mM DTT, 50 μM ATP, Substrate: Poly EY, 400 ng/μL, and recombinant Syk: variable.

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Kinase Assay - DELFIA Image 2

Figure 3. Dose dependence curve of Syk kinase activity: DELFIA® data generated using Phospho-Tyrosine Mouse mAb (P-Tyr-100) #9411 to detect phosphorylation of substrate peptide (#1310) by Syk kinase. In a 50 µl reaction, increasing amounts of Syk and 1.5 µM substrate peptide were used per reaction at room temperature for 30 minutes. (DELFIA® is a registered trademark of PerkinElmer, Inc.)

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Kinase Assay - DELFIA Image 3

Figure 5. Staurosporine inhibition of Syk kinase activity: DELFIA® data generated using Phospho-Tyrosine Mouse mAb (P-Tyr-100) #9411 to detect phosphorylation of Syk substrate peptide (#1310) by Syk kinase. In a 50 µl reaction, 50 ng Syk, 1.5 µM substrate peptide, 20 µM ATP and increasing amounts of staurosporine were used per reaction at room temperature for 30 minutes. (DELFIA® is a registered trademark of PerkinElmer, Inc.)

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Kinase Assay - DELFIA Image 4

Figure 4. Peptide concentration dependence of Syk kinase activity: DELFIA® data generated using Phospho-Tyrosine Mouse mAb (P-Tyr-100) #9411 to detect phosphorylation of substrate peptide (#1310) by Syk kinase. In a 50 µl reaction, 50 ng of Syk and increasing concentrations of substrate peptide were used per reaction at room temperature for 30 minutes. (DELFIA® is a registered trademark of PerkinElmer, Inc.)

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Kinase Assay - DELFIA Image 5

Figure 2. Time course of Syk kinase activity: DELFIA® data generated using Phospho-Tyrosine Mouse mAb (P-Tyr-100) #9411 to detect phosphorylation of Syk substrate peptide (#1310) by Syk kinase. In a 50 µl reaction, 50 ng Syk and 1.5 µM substrate peptide were used per reaction. (DELFIA® is a registered trademark of PerkinElmer, Inc.)

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Product Includes Quantity (with Count) Solution Color
DTT (1000X) 1 x 80 µl
ATP (10 mM) 9804 1 x 1 ml

The kit provides a means of performing kinase activity assays with recombinant human Syk kinase. It includes active Syk kinase (supplied as a GST fusion protein), a biotinylated peptide substrate and a phospho-tyrosine antibody for detection of the phosphorylated form of the substrate peptide.

Molecular Formula:

Peptide substrate, Biotin-Gastrin Precursor (Tyr87): 2,853 Daltons. GST-Syk Kinase: 104 kDa

Peptide Core Sequence:

EAY*GW

Syk is a protein tyrosine kinase that plays an important role in intracellular signal transduction in hematopoietic cells (1-3). Syk interacts with immunoreceptor tyrosine-based activation motifs (ITAMs) located in the cytoplasmic domains of immune receptors (4). It couples the activated immunoreceptors to downstream signaling events that mediate diverse cellular responses, including proliferation, differentiation, and phagocytosis (4). There is also evidence of a role for Syk in nonimmune cells and investigators have indicated that Syk is a potential tumor suppressor in human breast carcinomas (5). Tyr323 is a negative regulatory phosphorylation site within the SH2-kinase linker region in Syk. Phosphorylation at Tyr323 provides a direct binding site for the TKB domain of Cbl (6,7). Tyr352 of Syk is involved in the association of PLCγ1 (8). Tyr525 and Tyr526 are located in the activation loop of the Syk kinase domain; phosphorylation at Tyr525/526 of human Syk (equivalent to Tyr519/520 of mouse Syk) is essential for Syk function (9).

  1. Cheng, A.M. and Chan, A.C. (1997) Curr Opin Immunol 9, 528-33.
  2. Kurosaki, T. (1997) Curr Opin Immunol 9, 309-18.
  3. Chu, D.H. et al. (1998) Immunol Rev 165, 167-80.
  4. Turner, M. et al. (2000) Immunol Today 21, 148-54.
  5. Coopman, P.J. et al. (2000) Nature 406, 742-7.
  6. Deckert, M. et al. (1998) J Biol Chem 273, 8867-74.
  7. Rao, N. et al. (2001) EMBO J 20, 7085-95.
  8. Law, C.L. et al. (1996) Mol Cell Biol 16, 1305-15.
  9. Zhang, J. et al. (2000) J Biol Chem 275, 35442-7.
Entrez-Gene Id
6850
Swiss-Prot Acc.
P43405
For Research Use Only. Not For Use In Diagnostic Procedures.

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
DELFIA is a registered trademark of PerkinElmer, Inc.

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