80801 Gαs Activation Assay Kit_详细资料

产品信息

Product Description
A structurally diverse repertoire of ligands, from photons to large peptides, activates G protein-coupled
receptors （GPCRs） to elicit their physiological functions. Ligand-bound GPCRs, in turn, function as
guanine nucleotide exchange factors catalyzing the exchange of GDP bound on the Gα subunit with
GTP in the presence of Gβγ, causing the dissociation of the Gα subunit from the Gβγ dimer to form
two functional units （Gα and Gβγ）. Both Gα and Gβγ subunits signal to various cellular signaling
pathways. Based on the sequence and functional homologies, G proteins are grouped into four families:
Gs, Gi, Gq, and G12.
Gαs family relays signals from many GPCRs to regualte various biological functions such as the
stimulation of adenylyl cyclases. There were no direct methods to measure the activation of Gαs
proteins by receptors （until this assay kit）. Most reports used one downstream pathway, the increase
of cAMP, as a readout.
NewEast Biosciences Gαs Activation Assay Kit provides a direct measurement of the activation of Gαs
proteins. This is a simple and fast tool to monitor the activation of Gαs. Each kit provides sufficient
quantities to perform 20 assays.
NewEast Biosciences Gαs Activation Assay Kit is based on the monoclonal antibody specifically
recognizing the active GTP-bound Gαs proteins. This monoclonal antibody has much lower affinity
towards the inactive Gαs proteins. Therefore, after activation by receptor signals, active GTP-bound
Gαs proteins could be immunoprecipitated by this monoclonal antibody and further quantified by
western blot with another anti- Gαs antibody.
Assay Principle
NewEast Biosciences Gαs Activation Assay Kit is an immunoprecipitation/western blot assay to
measure the levels of active GTP-bound Gαs proteins, either from cell extracts or from in vitro GTPγS
loaded Gαs proteins. Briefly, the anti-active Gαs monoclonal antibody will specifically bind to active
Gαs protein. This antibody/Gαs complex will then be pulled down by protein A/G agarose. The
precipitated active Gαs proteins will be detected by immunoblots with another anti-Gαs antibody.
Kit Components
1. Anti-active Gαs, Mouse Monoclonal Antibody （Catalog No. 26906）: One vial – 22 μL （1
mg/mL） in PBS, pH 7.4, contained 50% glycerol. This antibody specifically recognizes
GTP- Gαs 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. 30303）: One bottle – 30 mL of 250 mM Tris-HCl, pH 7.4,
750 mM NaCl, 5 mM EDTA, 5% Triton X-100.
4. Anti-Gαs, Mouse Monoclonal Antibody （Catalog No. 26006）: One vial – 22 μL（1 mg/mL） in
PBS, pH 7.4, contained 50% glycerol.
5. 100 X GTPγS （Catalog No. 30302）: 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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