This volume explores techniques used to study either the structure or the functions of caveolae and their components in several normal and pathophysiological situations. The chapters in this book cover topics such as selective visualization of caveolae by electron microscopy techniques; spatiotemporal analysis of caveolae dynamics and mechanics using live cell fluorescence microscopy; in vitro reconstitution such as liposomes and GPMVs as tools to study caveolin- and cavin-interacting partners; pulling of tethers from the cell plasma membrane using optical tweezers; and immunofluorescence-based analysis of caveolin-3 in the diagnostic management of neuromuscular diseases. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls.
Authoritative and comprehensive, Caveolae: Methods and Protocols is a valuable resource for both novice and expert researchers who are interested in discovering new roles or regulations of formed caveolae, and proteins composing their coat in various model organisms.
Selective Visualization of Caveolae by TEM using APEX2.- Freeze-Fracture
Replica Immunolabeling of Cryopreserved Membrane Compartments, Cultured Cells
and Tissues.- Analysis of Caveolin in Primary Cilia.- Method for Efficient
Observation of Caveolin-1 Plasma Membrane by Microscopy Imaging
Analysis.- Quantitative Image Analysis of the Spatial Organization and
Mobility of Caveolin Aggregates at the Plasma Membrane.- Spatiotemporal
Analysis of Caveolae Dynamics using Total Internal Reflection Fluorescence
Microscopy.- Live-Cell FRET Imaging of Phosphorylation-Dependent Caveolin-1
Switch.- GPMVs as a Tool to Study Caveolin-Interacting Partners.- Biotin
Proximity Labeling to Identify Protein-Protein Interactions for
Cavin1.- Investigation of Novel Cavin-1/Suppressor of Cytokine 3 (SOCS3)
Interactions by Co-Immunoprecipitation, Peptide Pull-Down and Peptide Array
Overlay Approaches.- Analysis of Protein and Lipid Interactions using
Liposome Co-Sedimentation Assays.- Liposome Binding Assay to Characterize
Cavin Family Protein Structure and Functions.- Preparation of Caveolin-1 for
NMR Spectroscopy Experiments.- Tagging and Deleting of Endogenous Caveolar
Components using CRISPR/Cas9 Technology.- Pulling of Tethers from the Cell
Plasma Membrane using Optical Tweezers.- Live Confocal Imaging of Zebrafish
Notochord Cells under Mechanical Stress In Vivo.- Study of Caveolae Dependent
Mechanoprotection in Huma Muscle Cells using Micropatterning and Live Cell
Microscopy.- Immunofluorescence-Based Analysis of Caveolin-3 in the
Diagnostic Management of Neuromuscular Diseases.
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