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7743
HTScan® FLT3 Kinase Assay Kit
Cellular Assay Kits
Assay Kit

HTScan® FLT3 Kinase Assay Kit #7743

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Figure 1. FLT3 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: MBP 200 ng/μL, and variable amounts of FLT3.
Figure 5. Staurosporine inhibition of FLT3 kinase activity: DELFIA® data generated using Phospho-Tyrosine mAb (P-Tyr-100) #9411 to detect phosphorylation of FLT3 substrate peptide (#1310) by FLT3 kinase. In a 50 µl reaction, 50 ng FLT3, 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.)
Figure 3. Dose dependence curve of FLT3 kinase activity: DELFIA® data generated using Phospho-Tyrosine mAb (P-Tyr-100) #9411 to detect phosphorylation of substrate peptide (#1310) by FLT3 kinase. In a 50 µl reaction, increasing amounts of FLT3 and 1.5 µM substrate peptide were used per reaction at room temperature for 30 minutes. (DELFIA® is a registered trademark of PerkinElmer, Inc.)
Figure 4. Peptide concentration dependence of FLT3 kinase activity: DELFIA® data generated using Phospho-Tyrosine mAb (P-Tyr-100) #9411 to detect phosphorylation of substrate peptide (#1310) by FLT3 kinase. In a 50 µl reaction, 50 ng of FLT3 and increasing concentrations of substrate peptide were used per reaction at room temperature for 30 minutes. (DELFIA® is a registered trademark of PerkinElmer, Inc.)
Figure 2. Time course of FLT3 kinase activity: DELFIA® data generated using Phospho-Tyrosine mAb (P-Tyr-100) #9411 to detect phosphorylation of FLT3 substrate peptide (#1310) by FLT3 kinase. In a 50 µl reaction, 50 ng FLT3 and 1.5 µM substrate peptide were used per reaction. (DELFIA® is a registered trademark of PerkinElmer, Inc.)
Inquiry Info.# 7743

Product Description

The kit provides a means of performing kinase activity assays with recombinant human FLT3 kinase. It includes active FLT3 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-peptide: 2,853 Daltons. GST-FLT3 Kinase: 82 kDa.

Background

FMS-related tyrosine kinase 3 (FLT3, also called FLK2) is a member of the Type III receptor tyrosine kinase family, which includes c-Kit, PDGFR, and M-CSF receptors. FLT3 is expressed on early hematopoietic progenitor cells and supports growth and differentiation within the hematopoietic system (1,2). FLT3 is activated after binding with its ligand FL, which results in a cascade of tyrosine autophosphorylation and tyrosine phosphorylation of downstream substrates (3). The p85 subunit of PI3 kinase, SHP2, GRB2, and Shc have all been reported to associate with FLT3 after FL stimulation (4-6). Tyr589/591 is located in the juxtamembrane region of FLT3 and may play an important role in regulation of FLT3 tyrosine kinase activity. Somatic mutations of FLT3 consisting of internal tandem duplications (ITDs) occur in 20% of patients with acute myeloid leukemia (7).

Pathways

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