80701 Cdc42 Activation Assay Kit_详细资料

产品信息

Product Description
Small GTPases are a super-family of cellular signaling regulators. Cdc42 belongs to the Rho sub-family
of GTPases that regulate cell motility, cell division, and gene transcription. GTP binding increases the
activity of Cdc42, and the hydrolysis of GTP to GDP renders it inactive.
Currently the activation of Cdc42 proteins is assayed with the binding of GTP-bound Cdc42 to the
p21-binding domain （PBD） of p21-activated protein kinase （PAK）. This method is based on the
observation that the active, GTP-bound Cdc42 （and Rac） could bind to the PBD of PAK. However, the
reproducibility of this method is poor. This is partially due to the relatively quick hydrolysis of GTP to
GDP during the assay procedure, and the low binding affinity of PBD to Cdc42-GTP.
NewEast Biosciences Cdc42 Activation Assay Kit is based on the configuration-specific monoclonal
antibody that specifically recognizes Cdc42-GTP, but not Cdc42-GDP. Given the high affinity of
monoclonal antibodies to their antigens, the activation assay could be performed in a much shorter time.
This assay provides the reliable results with consistent reproducibility.
These anti-Cdc42-GTP monoclonal antibody can also be used to monitor the activation of Cdc42 in
cells and in tissues by immunohistochemistry.
NewEast Biosciences Cdc42 Activation Assay Kit provides a simple and fast method to monitor the
activation of Cdc42. Each kit provides sufficient quantities to perform 20 assays.
Assay Principle
NewEast Biosciences bases on the configuration-specific anti-Cdc42-GTP
monoclonal antibody to measure the active Cdc42-GTP levels, either from cell extracts or from in vitro
GTPγS loading Cdc42 activation assays. Briefly, anti-active Cdc42 mouse monoclonal antibody will
be incubated with cell lysates containing Cdc42-GTP. The bound active Cdc42 will then be pulled
down by protein A/G agarose. The precipitated active Cdc42 will be detected by immunoblot analysis
using anti-Cdc42 rabbit polyclonal antibody.
Kit Components
1. Anti-active Cdc42, Mouse Monoclonal Antibody （Catalog No. 26905）: One vial – 22 μL （1
mg/ml） in PBS, pH 7.4, containing 50% glycerol and 0.05% sodium azide. This antibody
specifically recognizes Cdc42-GTP from all vertebrates.
2. Protein A/G Agarose （Catalog No. 30301）: One vial – 400 μL of 50% slurry.
3. 5X Assay/Lysis Buffer （Catalog No. 30302）: One bottle – 30 mL of 250 mM Tris-HCl, pH 8, 750
mM NaCl, 50 mM MgCl2, 5 mM EDTA, 5% Triton X-100.
4. Anti-Cdc42, Rabbit Polyclonal Antibody （Catalog No. 21010）: One vial – 22 μL （1 mg/ml） in
PBS, pH 7.4, contained 50% glycerol.
5. 100 X GTPγS （Catalog No. 30303）: One vial –100 μl at 10 mM, use 5 μL of GTPγS for
GTP-labeling of 0.5 mL of cell lysate.
6. 100 X GDP （Catalog No. 30304）: One vial –100 μl at 100 mM, use 5 μL of GDP for
GDP-labeling of 0.5 mL of cell lysate.

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