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7758
HTScan® Lck Kinase Assay Kit
Cellular Assay Kits
Assay Kit

HTScan® Lck Kinase Assay Kit #7758

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HTScan® Lck Kinase Assay Kit: Image 1

Figure 3. Dose dependence curve of Lck kinase activity: DELFIA® data generated using Phospho-Tyrosine mAb (P-Tyr-100) #9411 to detect phosphorylation of substrate peptide #1365 by Lck kinase. In a 50 µl reaction, increasing amounts of Lck and 1.5 µM substrate peptide were used per reaction well at 25ºC for 30 minutes. (DELFIA® is a registered trademark of PerkinElmer, Inc.)

HTScan® Lck Kinase Assay Kit: Image 2

Figure 5. Staurosporine inhibition of Lck kinase activity: DELFIA® data generated using Phospho-Tyrosine mAb (P-Tyr-100) #9411 to detect phosphorylation of Lck substrate peptide (#1365) by Lck kinase. In a 50 µl reaction, 50 ng Lck, 1.5 µM substrate peptide, 50 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.)

HTScan® Lck Kinase Assay Kit: Image 3

Figure 4. Peptide concentration dependence of Lck kinase activity: DELFIA® data generated using Phospho-Tyrosine mAb (P-Tyr-100) #9411 to detect phosphorylation of substrate peptide (#1365) by Lck kinase. In a 50 µl reaction, 50 ng of Lck and increasing concentrations of substrate peptide were used per reaction at room temperature for 30 minutes. (DELFIA® is a registered trademark of PerkinElmer, Inc.)

HTScan® Lck Kinase Assay Kit: Image 4

Figure 2. Time course of Lck kinase activity: DELFIA® data generated using Phospho-Tyrosine mAb (P-Tyr-100) #9411 to detect phosphorylation of Lck substrate peptide (#1365) by Lck kinase. In a 50 µl reaction, 50 ng and 1.5 µM substrate peptide were used per reaction. (DELFIA® is a registered trademark of PerkinElmer, Inc.)

HTScan® Lck Kinase Assay Kit: Image 5

Figure 1. Lck 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, 5 mM MgCl2, 0.05 mM DTT, 50 μM ATP, Substrate: Poly (EY 4:1) 400 ng/μL, and variable amount of Lck kinase.

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Product Description

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

Molecular Formula

Peptide substrate, Biotin-FHIT (Tyr114): 2,150 Daltons, GST-Lck Kinase: 84 kDa.

Background

The Src family of protein tyrosine kinases, which includes Src, Lyn, Fyn, Yes, Lck, Blk, and Hck, are important in the regulation of growth and differentiation of eukaryotic cells (1). Src activity is regulated by tyrosine phosphorylation at two sites, but with opposing effects. While phosphorylation at Tyr416 in the activation loop of the kinase domain upregulates enzyme activity, phosphorylation at Tyr527 in the carboxy-terminal tail by Csk renders the enzyme less active (2).

Lck is essential for T-lymphocyte activation and differentiation (3,4). Phosphorylation of Tyr505 in the carboxy-terminal tail of Lck downregulates its catalytic activity, while phosphorylation of Tyr394 leads to an increase in Lck activity (5).

  1. Thomas, S.M. and Brugge, J.S. (1997) Annu Rev Cell Dev Biol 13, 513-609.
  2. Hunter, T. (1987) Cell 49, 1-4.
  3. Molina, T.J. et al. (1992) Nature 357, 161-4.
  4. Straus, D.B. and Weiss, A. (1992) Cell 70, 585-93.
  5. Chow, L.M. et al. (1993) Nature 365, 156-60.

Pathways & Proteins

Explore pathways + proteins related to this product.

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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.