Receptor Signal Transduction Protocols

Inducible Expression of G Protein-Coupled Receptors in Transfected Cells.- Using the Flp-InTM T-RexTM System to Regulate GPCR Expression.- Viral Infection for GPCR Expression in Eukaryotic Cells.- Generation of Epitope-Tagged GPCRs.- The Use of Site-Directed Mutagenesis to Study GPCRs.- Approaches to Study GPCR Regulation in Native Systems.- Heterologous Expression of GPCRs in Fission Yeast.- Radioligand Binding Methods for Membrane Preparations and Intact Cells.- Quantification of GPCR mRNA Using Real-Time RT-PCR.- Determining Allosteric Modulator Mechanism of Action: Integration of Radioligand Binding and Functional Assay Data.- Design and Use of Fluorescent Ligands to Study Ligand-Receptor Interactions in Single Living Cells.- Examining Site-Specific GPCR Phosphorylation.- Ubiquitination of GPCRs.- [35S]GTPγS Binding as an Index of Total G-Protein and Gα-Subtype-Specific Activation by GPCRs.- Using Calcium Imaging as a Readout of GPCR Activation.- Measuring Spatio-Temporal Dynamics of Cyclic AMP Signaling in Real-Time Using FRET-Based Biosensors.- Determining the Activation of Rho as an Index of Receptor Coupling to G12/13 Proteins.- The Use of Translocating Fluorescent Biosensors for Real-Time Monitoring of GPCR-Mediated Signaling Events.- Study of GPCR-Protein Interactions Using Gel Overlay Assays and Glutathione-S-Transferase-Fusion Protein Pull-Downs.- Study of GPCR-Protein Interactions by BRET.- Time Resolved FRET Strategy with Fluorescent Ligands to Analyze Receptor Interactions in Native Tissues: Application to GPCR Oligomerization.- Peptide Affinity Purification for the Isolation and Identification of GPCR-AssociatedProtein Complexes.- Tandem Affinity Purification and Identification of GPCR-Associated Protein Complexes.- Identification of GPCR Localization in Detergent Resistant Membranes.- Analysis of GPCR Localization and Trafficking.- Statistical Methods in Research.