80301 Configuration-specific Monoclonal A...

Configuration-specific Monoclonal Antibody Based
Gαi Activation Assay Kit
Catalog Number： 80301
20 assays

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.
Gαi family is the largest family of G proteins. They relay signals from many GPCRs to regualte
various biological functions. There were no direct methods to measure the activation of Gαi proteins
by receptors （until this assay kit）. Most reports used one of the downstream pathway, i.e. the
inhibition of adenylyl cyclases, as a readout. Alternatively, sensitivity to pertussis toxin （PTX） was
used as an indicator of possible Gαi proteins invovled in a signaling pathway.
NewEast Biosciences Gαi Activation Assay Kit provides a direct measurement of the activation of Gαi
proteins. This is a simple and fast tool to monitor the activation of Gαi. Each kit provides sufficient
quantities to perform 20 assays.
NewEast Biosciences Gαi Activation Assay Kit is based on the monoclonal antibody specifically
recognizing the active GTP-bound Gαi proteins. This monoclonal antibody has much lower affinity
towards the inactive Gαi proteins. Therefore, after activation by receptor signals, active GTP-bound
Gαi proteins could be immunoprecipitated by this monoclonal antibody and further quantified by
western blot with another anti- Gαi antibody.
Assay Principle
NewEast Biosciences Gαi Activation Assay Kit is an immunoprecipitation/western blot assay to
measure the levels of active GTP-bound Gαi proteins, either from cell extracts or from in vitro GTPγS
loaded Gαi proteins. Briefly, the anti-active Gαi monoclonal antibody will specifically bind to active
Gαi protein. This antibody/ Gαi complex will then be pulled down by protein A/G agarose. The
precipitated active Gαi proteins will be detected by immunoblots with another anti-Gαi antibody.