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7701
HTScan® ABL1 Kinase Assay Kit
Cellular Assay Kits

HTScan® ABL1 Kinase Assay Kit #7701

This product is discontinued

Kinase Assay - Radiometric Image 1

Figure 5. ABL1 kinase activity was measured in a radioisotopic filtration assay using the following reaction conditions: 60 mM HEPES-NaOH, pH 7.5, 3 mM MgCl2, 3 mM MnCl2, 3 uM Na-orthovanadate, 1.2 mM DTT, ATP (variable), 2.5 ug / 50 ul PEG20.000, Substrate: PolyAEKY, 5 ug / 50 ul, Recombinant ABL1: 5 Units / 50 µl.

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Kinase Assay - DELFIA Image 2

Figure 3. Dose dependence curve of ABL1 kinase activity: DELFIA® data generated using Phospho-Tyrosine Monoclonal Antibody P-Tyr-100 #9411 to detect phosphorylation of peptide C03-719 by GST-ABL1 kinase. In a 50 ul reaction, increasing amounts of GST-ABL1 and 1.5 uM substrate peptide were used per reaction well at 25ºC for 30 minutes. Background reading is 1481. (DELFIA® is a registered trademark of PerkinElmer, Inc.)

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Kinase Assay - DELFIA Image 3

Figure 6. Staurosporine inhibition of ABL1 kinase activity: DELFIA® data generated using Phospho-Tyrosine Monoclonal Antibody P-Tyr-100 #9411 to detect phosphorylation of ABL1 substrate peptide (C03-719) by GST-ABL1 kinase. In a 50 ul reaction, 10 Units GST-ABL1 , 1.5 uM substrate peptide, 5 uM ATP and increasing amount of staurosporine were used per reaction well at 25ºC 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 ABL1 kinase activity: DELFIA® data generated using Phospho-Tyrosine Monoclonal Antibody P-Tyr-100 #9411 to detect phosphorylation of peptide C03-719 by GST-ABL1 kinase. In a 50 ul reaction, 10 Units of GST-ABL1 and increasing concentrations of substrate peptide were used per reaction well at 25ºC for 30 minutes. Background reading is 871. (DELFIA® is a registered trademark of PerkinElmer, Inc.)

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Kinase Assay - DELFIA Image 5

Figure 1. Time course of ABL1 kinase activity: DELFIA® data generated using Phospho-Tyrosine Monoclonal Antibody P-Tyr-100 #9411 to detect phosphorylation of ABL1 substrate peptide by GST-ABL1 kinase. In a 50 ul reaction, 10 Units GST-ABL1 and 1.5 uM substrate peptide were used per reaction well. Background reading is 768. (DELFIA® is a registered trademark of PerkinElmer, Inc.)

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Kinase Assay - DELFIA Image 6

Figure 2. Treatment of Tyrosine Kinase Substrate Screning Kit #7450 with GST-ABL1 Kinase. Positive control is in yellow and negative control is in blue. Several positive hits for ABL1 were identified using this approach (in red). The star indicates the peptide chosen for this HTScan® ABL1 Kinase Assay Kit.

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The kit provides a means of performing enzymatic assays with active human ABL1 kinase. It includes active ABL1 kinase (supplied as a GST fusion protein), a biotinylated substrate peptide and a phospho-tyorsine monoclonal antibody for detection of the phosphorylated form of the substrate peptide.

Molecular Formula:

Peptide Substrate, Biotin-Signal Transduction Protein (Tyr160): 1830 Daltons, GST-ABL1 Kinase domain: 74,140 Daltons

Peptide Core Sequence:

GIY*DV

The c-Abl proto-oncogene encodes a nonreceptor protein tyrosine kinase that is ubiquitously expressed and highly conserved in metazoan evolution. c-Abl protein is distributed in both the nucleus and the cytoplasm of cells. It is implicated in regulating cell proliferation, differentiation, apoptosis, cell adhesion, and stress responses (1-3). c-Abl kinase activity is increased in vivo by diverse physiological stimuli including integrin activation; PDGF stimulation; and binding to c-Jun, Nck, and RFX1 (2,4). The in vivo mechanism for regulation of c-Abl kinase activity is not completely understood. Tyr245 is located in the linker region between the SH2 and catalytic domains. This positioning is conserved among Abl family members. Phosphorylation at Tyr245 is involved in the activation of c-Abl kinase (5). In addition, phosphorylation at Tyr412, which is located in the kinase activation loop of c-Abl, is required for kinase activity (6).

  1. Wang, J.Y. (2000) Oncogene 19, 5643-50.
  2. Van Etten, R.A. (1999) Trends Cell Biol 9, 179-86.
  3. Danial, N.N. and Rothman, P. (2000) Oncogene 19, 2523-31.
  4. Shaul, Y. (2000) Cell Death Differ 7, 10-6.
  5. Brasher, B.B. and Van Etten, R.A. (2000) J Biol Chem 275, 35631-7.
  6. Pluk, H. et al. (2002) Cell 108, 247-259.
Entrez-Gene Id
25
Swiss-Prot Acc.
P00519
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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