G Protein-Coupled Receptors. Part A /

Mostrar otras versiones (1)

"The largest family of cell surface receptors in our body, referred to as G protein-coupled receptors, plays central roles in the onset and regulation of many different physiological processes. The cascade starts with binding of the "stimulus" (i.e., an activating ligand; agonist) to...

Descripción completa

Detalles Bibliográficos
Clasificación:Libro Electrónico
Otros Autores: Shukla, Arun K. (Editor )
Formato: Electrónico eBook
Idioma:Inglés
Publicado: Cambridge, MA, USA : Academic Press is an imprint of Elsevier, [2017]
Edición:Second edition.
Colección:Methods in cell biology ; v. 142.
Temas:
Cytology > Technique.
Cell receptors.
G proteins.
GTP-Binding Proteins
Cytological Techniques
Receptors, Cell Surface
Cytologie > Technique.
R�ecepteurs cellulaires.
Prot�eines G.
SCIENCE > Life Sciences > Anatomy & Physiology.
G proteins
Cell receptors
Cytology > Technique
Internet Resources.
Acceso en línea:Texto completo
Tabla de Contenidos:
  • 3.3. Transfection of Cells With the G[gamma] Translocation Sensor Using the Cationic Polymer, PEI3.3.1. Preparation of PEI; 3.3.2. Transfection; 3.3.3. Other methods for transfection; 3.4. Live-Cell Time-Lapse Imaging; 3.4.1. Examination of optical activation of light-sensing GPCRs (Opsins); 3.5. Image Analysis; 3.5.1. Manual processing; 3.5.2. Automated/Thresholding processing; 3.6. Stable Cell Line Generation; References; Chapter 2: GloSensor assay for discovery of GPCR-selective ligands; 1. Introduction; 2. cAMP Measurement Assays; 2.1. Fluorescence-Based cAMP Measuring Assays
  • Front Cover; G Protein-Coupled Receptors Part A; Copyright; Contents; Contributors; Preface; Chapter 1: Measurement of GPCR-G protein activity in living cells; 1. Introduction; 2. Materials; 2.1. Generations of Sensors; 2.2. Cell Culture; 2.3. Transfection of Cells; 2.4. Stable Cell Line Generation; 2.5. Live-Cell Imaging; 2.6. Fluorescence Imaging Plate Reader Assay; 3. Methods; 3.1. Generations of Sensors Using Gibson Assembly Protocol; 3.2. Cell Culture; 3.2.1. HeLa Cell-Specific Growth Media Preparation; 3.2.2. Passaging cells
  • 2.1.1. ALPHAscreen (Amplified Luminescent Proximity Homogenous Assay)2.1.2. DELFIA (Dissociation-Enhanced Lanthanide Fluorescence Immunoassay); 2.1.3. HTRF (Homogenous Time Resolved Fluorescence); 2.1.4. Fluorescence polarization assay; 2.2. Luminescence-Based cAMP Measuring Assays; 2.2.1. CRE-luciferase reporter gene assay; 2.2.2. GloSensor assay; 3. Reagents and Consumables; 4. Methods; 4.1. Cell Culture Maintenance and Trypsinization; 4.2. Transient Transfection of HEK293T Cells; 4.3. Preparation of Transfected Cell Plate; 4.4. Test Compound Dilution for GloSensor Assay
  • 4.5. Test Compound Addition to the Cell Plate4.6. Compound Addition to Screen Ligands for GS-Coupled Receptors; 4.7. Compound Addition to Screen Ligands for Gi-Coupled Receptors; 5. Data Analysis; 5.1. Data Analysis for Agonist Activity; 5.2. Data Analysis for Antagonist Activity; 6. Results and Data Interpretation; 7. Final Comments; Acknowledgments; References; Chapter 3: cAMP assays in GPCR drug discovery; 1. Introduction; 2. Materials; 2.1. Cell Culture; 3. Method; 3.1. Troubleshooting; 3.2. Notes; References
  • Chapter 4: Measurement of intracellular Ca2+ mobilization to study GPCR signal transduction1. Introduction; 2. Materials; 2.1. Maintenance of Cell Lines; 2.2. Generation of "Stable Cell Lines"; 2.3. Fura2AM Loading and Spectrophotometric Analysis; 3. Methods; 3.1. Maintenance of Cell Lines; 3.2. Generation of "Stable Cell Lines"; 3.3. Fura2AM Loading; 3.4. Spectrophotometric Analysis; 4. Notes; References; Chapter 5: Visualization and quantification of GPCR trafficking in mammalian cells by confocal microscopy; 1. Introduction; 2. Methods; 2.1. Required Materials

Ejemplares similares