Revision 1
Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
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UniProt ID:

#P35916

Entrez-Gene Id:

2324

Product Information

Storage

Antibodies are supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA and 50% glycerol. Do not aliquot the antibodies. Peptides are supplied at 6 µM in 0.001% DMSO. Enzymes are supplied in 50 mM Tris-HCL (pH 8.0), 100 mM NaCl, 5 mM DTT, 15 mM reduced glutathione and 20%
glycerol. Store at –80°C.
Keep enzymes on ice during use.
Avoid repeated freeze-thaw cycles.

Source / Purification

The GST-Kinase fusion protein was produced using a baculovirus expression system with a construct expressing a fragment of human VEGFR-3 (Asn799-Arg1298) (GenBank accession No. NM_002020) with an amino-terminal GST tag. The protein was purified by one-step affinity chromatography using glutathione-agarose.

Product Description

The kit provides a means of performing kinase activity assays with recombinant human VEGFR-3 kinase. It includes active VEGFR-3 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-MET (Tyr1253) peptide: 2,200Daltons. GST-VEGFR-3 Kinase: 86 kDa.

Background

Vascular endothelial growth factor receptor 3 (VEGFR3) is a 195 kDa membrane receptor tyrosine kinase. VEGF receptors are characterized by the presence of seven extracellular immunoglobulin (Ig)-like domains followed by a membrane-spanning domain and a conserved intracellular tyrosine kinase domain (1). VEGF receptor 3 expression is largely restricted to adult lymphatic endothelium and is thought to control lymphangiogenesis (1,2). Binding of VEGF-C/VEGF-D to VEGFR3 results in transphosphorylation of tyrosine residues in its intracellular domain, recruitment of signaling molecules and activation of ERK1/2 and Akt signaling cascades (1,3).

  1. Robinson, C.J. and Stringer, S.E. (2001) J. Cell Sci.114, 853-865.
  2. Valtola, R. et al. (1999) Am. J. Pathol. 154, 1381-1390.
  3. Saharinen, P. and Petrova, T.V. (2004) Ann. N.Y. Acad. Sci. 1014, 76-87.

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Cross-Reactivity Key

H: human M: mouse R: rat Hm: hamster Mk: monkey Vir: virus Mi: mink C: chicken Dm: D. melanogaster X: Xenopus Z: zebrafish B: bovine Dg: dog Pg: pig Sc: S. cerevisiae Ce: C. elegans Hr: horse GP: Guinea Pig Rab: rabbit All: all species expected

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

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Orders: 877-616-CELL (2355)•[email protected]• Support: 877-678-TECH (8324)•[email protected]• Web: cellsignal.com
仅供研究使用。不得用于诊断流程。
Revision 1
#7791

HTScan® VEGF Receptor 3 Kinase Assay Kit

HTScan® VEGF Receptor 3 Kinase Assay Kit: Image 1 Expand Image
图 3. Dose dependence curve of VEGFR-3 kinase activity: DELFIA® data generated using Phospho-Tyrosine Mouse mAb (P-Tyr-100) #9411 to detect phosphorylation of substrate peptide (#1367) by VEGFR-3 kinase. 在 50 µl 反应体积中，每个反应孔在室温下用数量递增的 VEGFR-3 和 1.5 µM 底物肽反应 30 分钟。（DELFIA® 是 PerkinElmer, Inc. 的注册商标）
HTScan® VEGF Receptor 3 Kinase Assay Kit: Image 2 Expand Image
图 5. Staurosporine inhibition of VEGFR-3 kinase activity: DELFIA® data generated using Phospho-Tyrosine Mouse mAb (P-Tyr-100) #9411 to detect phosphorylation of VEGFR-3 substrate peptide (#1367) by VEGFR-3 kinase. 在 50 µl 反应体积中，每个反应孔在室温下用 100 ng VEGFR-3、1.5 µM 底物肽、20 µM ATP 和数量递增的 staurosporine 反应 30 分钟。（DELFIA® 是 PerkinElmer, Inc. 的注册商标）
HTScan® VEGF Receptor 3 Kinase Assay Kit: Image 3 Expand Image
图 4. Peptide concentration dependence of VEGFR-3 kinase activity: DELFIA® data generated using Phospho-Tyrosine Mouse mAb (P-Tyr-100) #9411 to detect phosphorylation of substrate peptide (#1367) by VEGFR-3 kinase. 在 50 µl 反应体积中，每个反应孔在室温下用 100 ng VEGFR-3 和浓度递增的底物肽反应 30 分钟。（DELFIA® 是 PerkinElmer, Inc. 的注册商标）
HTScan® VEGF Receptor 3 Kinase Assay Kit: Image 4 Expand Image
图 2. Time course of VEGFR-3 kinase activity: DELFIA® data generated using Phospho-Tyrosine Mouse mAb (P-Tyr-100) #9411 to detect phosphorylation of VEGFR-3 substrate peptide (#1367) by VEGFR-3 kinase. 在 50 µl 反应体积中，每个反应孔用 100 ng VEGFR-3 和 1.5 µM 底物肽。（DELFIA® 是 PerkinElmer, Inc. 的注册商标）
HTScan® VEGF Receptor 3 Kinase Assay Kit: Image 5 Expand Image
图 1. 使用以下反应条件在放射测定法中测量 VEGFR2 激酶活性： 60 mM HEPES-NaOH、pH 7.5、3 mM MgCl2, 3 mM MnCl2, 3 µM 原钒酸钠、1.2 mM DTT、ATP（变量）、2.5 µg/50 µl PEG20,000，底物：PolyEY5 µg/50 µl、重组 VEGFR-3：100 ng/50 µl。
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Orders: 877-616-CELL (2355)•[email protected]• Support: 877-678-TECH (8324)•[email protected]• Web: cellsignal.com
仅供研究使用。不得用于诊断流程。